Not an inhibitor of alpha/beta hydrolases. Glutathione depleting agent. Used to test the involvment of glutathione-S-transferases in the resistance to insecticides. One of the key uses for the compound is in production of the pesticide Malathion
38 moreTitle: The overexpression of three cytochrome P450 genes CYP6CY14, CYP6CY22 and CYP6UN1 contributed to metabolic resistance to dinotefuran in melon/cotton aphid, Aphis gossypii Glover Chen A, Zhang H, Shan T, Shi X, Gao X Ref: Pestic Biochem Physiol, 167:104601, 2020 : PubMed
Dinotefuran, the third-generation neonicotinoid, has been applied against melon/cotton aphid Aphis gossypii Glover in China. The risk of resistance development, cross-resistance pattern and potential resistance mechanism of dinotefuran in A. gossypii were investigated. A dinotefuran-resistant strain of A. gossypii (DinR) with 74.7-fold resistance was established by continuous selection using dinotefuran. The DinR strain showed a medium level of cross resistance to thiamethoxam (15.2-fold), but no cross resistance to imidacloprid. The synergism assay indicated that piperonyl butoxide and triphenyl phosphate showed synergistic effects on dinotefuran toxicity to the DinR strain with a synergistic ratio of 8.3 and 2.5, respectively, while diethyl maleate showed no synergistic effect. The activities of cytochrome P450 monooxygenase and carboxylesterase were significantly higher in DinR strain than in susceptible strain (SS). Moreover, the gene expression results showed that CYP6CY14, CYP6CY22 and CYP6UN1 were significantly overexpressed in DinR strain compared with SS strain. The expression of CYP6CY14 was 5.8-fold higher in DinR strain than in SS strain. Additionally, the transcription of CYP6CY14, CYP6CY22 and CYP6UN1 in A. gossypii showed dose- and time-dependent responses to dinotefuran exposure. Furthermore, knockdown of CYP6CY14, CYP6CY22 and CYP6UN1 via RNA interference (RNAi) significantly increased mortality of A. gossypii, when A. gossypii was treated with dinotefuran. These results demonstrated the overexpression of CYP6CY14, CYP6CY22 and CYP6UN1 contributed to dinotefuran resistance in A. gossypii.
        
Title: Synergism of organophosphorus insecticides by diethyl maleate and related compounds in house flies Welling W, de Vries JW Ref: Pesticide Biochemistry and Physiology, 23:358, 1985 : PubMed
The toxicity of several organophosphorus and one carbamate insecticide for house flies is enhanced by simultaneous administration of diethyl maleate. Synergism factors vary from 2 to 116 and are strongly dependent on the combination of strain and insecticide studied. In general, it seems that thiono compounds are less synergized than their oxon analogs. Comparison of some analogs of maleic acid esters showed that the diethyl ester was not the most active compound. trans-Phenylbutenone was found to be a good alternative for diethyl maleate. Both diethyl maleate and trans-phenylbutenone deplete glutathione in the flies, but this depletion per se is certainly not the most important mode of action. In vitro experiments made clear that sufficient glutathione remained to permit a substantial insecticide degradation rate. Moreover, treatment of the flies with diethyl maleate 24 hr before the insecticide application resulted in a partial or complete disappearance of synergism in a period in which the glutathione concentration in the flies was still very low. Apart from this effect on the glutathione levels in the insect tissues it appeared from further experimental work in vitro that both compounds had a direct inhibitory effect on glutathione-dependent transferase(s) as well as on oxidative enzymes involved in insecticide degradation. The contribution of these reactions to the eventual synergism factor is discussed.
        
Title: Parathion and methyl parathion toxicity and metabolism in piperonyl butoxide and diethyl maleate pretreated mice Mirer FE, Levine BS, Murphy SD Ref: Chemico-Biological Interactions, 17:99, 1977 : PubMed
38 lessTitle: Solvent-dependent activity of Candida antarctica lipase B and its correlation with a regioselective mono aza-Michael addition - experimental and molecular dynamics simulation studies Nazarian Z, Arab SS Ref: Heliyon, 8:e10336, 2022 : PubMed
With the aim of gaining understanding of the molecular basis of commercially available Candida antarctica lipase B (CALB) immobilized on polyacrylic resin catalyzed regioselective mono aza-Michael addition of Benzhydrazide to Diethyl maleate we decided to carry out molecular dynamics (MD) simulation studies in parallel with our experimental study. We found a correlation between the activity of CALB and the choice of solvent. Our study showed that solvent affects the performance of the enzyme due to the binding of solvent molecules to the enzyme active site region, and the solvation energy of substrates in the different solvents. We also found that CALB is only active in nonpolar solvent (i.e. Hexane), and therefore we investigated the influence of Hexane on the catalytic activity of CALB for the reaction. The results of this study and related experimental validation from our studies have been discussed here.
        
Title: Associations between acetylcholinesterase-1 mutations and chlorpyrifos resistance in beet armyworm, Spodoptera exigua Teng H, Zuo Y, Jin Z, Wu Y, Yang Y Ref: Pestic Biochem Physiol, 184:105105, 2022 : PubMed
Control of the beet armyworm, Spodoptera exigua depends heavily on chemical insecticides. Chlorpyrifos, an acetylcholinesterase (AChE) inhibitor, has been used in beet armyworm control for many years in China. Here we describe high level resistance to chlorpyrifos in a S. exigua strain, FX19-R, which was developed from a field-collected Chinese strain (FX) by selection with chlorpyrifos in the laboratory. FX19-R showed 1001-fold resistance to chlorpyrifos compared with the laboratory reference strain WH-S. The esterase inhibitor triphenyl phosphate (TPP) provided significant but small synergism (only 3.5-fold) for chlorpyrifos and neither of the glutathione s-transferase depletor diethyl maleate and the cytochrome P450s inhibitor piperonyl butoxide provided any detectable synergism, indicating that AChE insensitivity may play the major role in the resistance in FX19-R. Consistent with this, an amino acid substitution, F443Y (F331Y in standard Torpedo californica numbering) in AChE1 was identified in the FX19-R strain and shown to be tightly linked to chlorpyrifos resistance. Precisely homologous substitutions have been associated with organophosphate resistance in other pest species. A novel amino acid substitution, G311S (or G198S in standard numbering), was also identified in the reference strain WH-S. Recombinantly expressed AChE1 proteins carrying the G311S and F443Y substitutions were about 4.2-fold and 210-fold less sensitive to inhibition by chlorpyrifos oxon than wild-type AChE1, respectively. These results enhance our understanding of the mechanisms of chlorpyrifos resistance and provide a basis for resistance management based on monitoring the F443Y and G311S substitutions.
        
Title: Multiple Mechanisms Conferring Broad-Spectrum Insecticide Resistance in the Tropical Bed Bug (Hemiptera: Cimicidae) Dang K, Doggett SL, Leong XY, Veera Singham G, Lee CY Ref: J Econ Entomol, :, 2021 : PubMed
The modern resurgence of the common (Cimex lectularius L.) and tropical bed bugs (C. hemipterus [F.]) is thought to be primarily due to insecticide resistance. While there are many reports on insecticide resistance mechanisms in C. lectularius, such information in C. hemipterus is limited. We examined dichloro-diphenyl-trichloroethane (DDT), malathion, deltamethrin, permethrin, lambda-cyhalothrin resistance, and the underlying mechanisms in several C. hemipterus strains (Australia: Queensland [QLD-AU]; Malaysia: Kuala Lumpur [KL-MY], Tanjung Tokong [TT-MY], Christian [CH-MY], and Green Lane [GL-MY]). We used a surface contact method, synergism studies (utilizing piperonyl butoxide [PBO], S,S,S-tributyl phosphorotrithioate [DEF], and diethyl maleate [DEM]), and molecular detection of kdr mutations. Results demonstrated that all C. hemipterus strains possessed high resistance to DDT and the pyrethroids and moderate to high resistance to malathion. Synergism studies showed that deltamethrin resistance in all strains was significantly (P < 0.05) inhibited by PBO. In contrast, deltamethrin resistance was not affected in DEF or DEM. Similar findings were found with lambda-cyhalothrin resistance. Malathion resistance was significantly (P < 0.05) reduced by DEF in all strains. Resistance to DDT was not affected by DEM in all strains. Multiple kdr mutations (M918I, D953G, and L1014F) were detected by molecular analyses. TT-MY strain was found with individuals possessing three kdr mutation combinations; D953G + L1014F (homozygous susceptible: M918), M918I + D953G + L1014F (heterozygous resistant: I918), and M918I + D953G + L1014F (homozygous resistant: I918). Individuals with M918I + D953G + L1014F (homozygous resistant: I918) survived longer on deltamethrin (>12 h) than those (>=1 h) with other combinations. M918I + L1014F mutations most likely conferred super-kdr characteristic toward pyrethroids and DDT in C. hemipterus.
BACKGROUND: Fall armyworm (FAW), Spodoptera frugiperda (Smith), is an economically important pest worldwide. In this study, we selected a genotype of FAW resistant to chlorpyrifos from a field-collected population, characterized the genetic basis of resistance, evaluated cross-resistance and mechanisms of resistance using synergists. RESULTS: The LD(50) values of chlorpyrifos for the resistant (Clorp-R) and susceptible (Sus) FAW genotypes were 24.26 and 0.023 microg larva(-1) , respectively, representing a resistance ratio >1,050-fold. The LD(50) values of chlorpyrifos against heterozygotes were 3.34 and 4.00 microg larva(-1) , suggesting that resistance is autosomally inherited. The chlorpyrifos resistance in FAW was influenced by few genes, with the minimum numbers of segregations being 1.74 and 1.88. On chlorpyrifos-sprayed plants and leaves, Clorp-R and heterozygotes genotypes showed >95% and >52% survival, respectively, whereas the Sus genotype had no survival, indicating that the resistance is incompletely dominant at the field rate of chlorpyrifos. The Clorp-R genotype presented some cross-resistance to acephate, but low cross-resistance to thiodicarb, methomyl, chlorfenapyr, flubendiamide, methoxyfenozide, spinetoram, and teflubenzuron. The synergists piperonyl butoxide, diethyl maleate and S,S,S-tributyl phosphorotrithiotate did not have relevant effects on the Clorp-R genotype, suggesting a minor role for metabolic resistance. CONCLUSIONS: The inheritance of chlorpyrifos resistance in FAW was characterized as autosomal, incompletely dominant and polygenic, with metabolic resistance playing a small role in the detoxification of chlorpyrifos. Low cross-resistance between chlorpyrifos and other Mode of Action (MoA) insecticides occurs in FAW, highlighting the importance of considering the rotation of MoA as a strategy to delay resistance. This article is protected by copyright. All rights reserved.
        
Title: Metabolic Resistance in Permethrin-Resistant Florida Aedes aegypti (Diptera: Culicidae) Schluep SM, Buckner EA Ref: Insects, 12:, 2021 : PubMed
Aedes aegypti is the principal mosquito vector for many arthropod-borne viruses (arboviruses) including dengue, chikungunya, and Zika. In the United States, excessive permethrin use has led to a high frequency of resistance in mosquitoes. Insecticide resistance is a significant obstacle in the struggle against vector-borne diseases. To help overcome metabolic resistance, synergists that inhibit specific metabolic enzymes can be added to formulated pyrethroid products. Using modified CDC bottle bioassays, we assessed the effect of three inhibitors (piperonyl butoxide (PBO), which inhibits oxidase activity; S.S.S-tributyl phosphorotrithioate (DEF), which inhibits esterase activity; and diethyl maleate (DM), which inhibits glutathione transferase activity) + permethrin. We performed these against 20 Florida Ae. aegypti populations, all of which were resistant to permethrin. Our data indicated that 11 out of 20 populations (55%) exhibited metabolic resistance. Results revealed 73% of these populations had significant increases in mortality attributed to DEF + permethrin, 64% to PBO + permethrin, and 55% to DM + permethrin compared to permethrin alone. Currently, PBO is the only metabolic enzyme inhibitor added to formulated pyrethroid products used for adult mosquito control. Our results suggest that the DEF and DM inhibitors could also be useful additives in permethrin products, especially against metabolically resistant Ae. aegypti mosquitoes. Moreover, metabolic assays should be conducted to better inform mosquito control programs for designing and implementing integrated vector management strategies.
        
Title: The overexpression of three cytochrome P450 genes CYP6CY14, CYP6CY22 and CYP6UN1 contributed to metabolic resistance to dinotefuran in melon/cotton aphid, Aphis gossypii Glover Chen A, Zhang H, Shan T, Shi X, Gao X Ref: Pestic Biochem Physiol, 167:104601, 2020 : PubMed
Dinotefuran, the third-generation neonicotinoid, has been applied against melon/cotton aphid Aphis gossypii Glover in China. The risk of resistance development, cross-resistance pattern and potential resistance mechanism of dinotefuran in A. gossypii were investigated. A dinotefuran-resistant strain of A. gossypii (DinR) with 74.7-fold resistance was established by continuous selection using dinotefuran. The DinR strain showed a medium level of cross resistance to thiamethoxam (15.2-fold), but no cross resistance to imidacloprid. The synergism assay indicated that piperonyl butoxide and triphenyl phosphate showed synergistic effects on dinotefuran toxicity to the DinR strain with a synergistic ratio of 8.3 and 2.5, respectively, while diethyl maleate showed no synergistic effect. The activities of cytochrome P450 monooxygenase and carboxylesterase were significantly higher in DinR strain than in susceptible strain (SS). Moreover, the gene expression results showed that CYP6CY14, CYP6CY22 and CYP6UN1 were significantly overexpressed in DinR strain compared with SS strain. The expression of CYP6CY14 was 5.8-fold higher in DinR strain than in SS strain. Additionally, the transcription of CYP6CY14, CYP6CY22 and CYP6UN1 in A. gossypii showed dose- and time-dependent responses to dinotefuran exposure. Furthermore, knockdown of CYP6CY14, CYP6CY22 and CYP6UN1 via RNA interference (RNAi) significantly increased mortality of A. gossypii, when A. gossypii was treated with dinotefuran. These results demonstrated the overexpression of CYP6CY14, CYP6CY22 and CYP6UN1 contributed to dinotefuran resistance in A. gossypii.
        
Title: Biochemical Mechanisms, Cross-resistance and Stability of Resistance to Metaflumizone in Plutella xylostella Shen J, Li Z, Li D, Wang R, Zhang S, You H, Li J Ref: Insects, 11:, 2020 : PubMed
The diamondback moth, Plutella xylostella (L.) is an important pest of cruciferous crops worldwide. It has developed resistance to many conventional and novel insecticide classes. Metaflumizone belongs to the new chemical class of semicarbazone insecticides. To delay the development of metaflumizone resistance in P. xylostella and to guide insecticide use in the field, the biochemical mechanisms, cross-resistance spectrum, and stability of resistance to metaflumizone were studied in a laboratory-selected resistant strain (metaflu-SEL). Synergism tests with the carboxylesterase inhibitor triphenyl phosphate (TPP), the glutathione S-transferase depletor diethyl maleate (DEM), and the P450 inhibitor piperonyl butoxide(PBO) had no obvious effect on metaflumizone in the metaflu-SEL strain and the susceptible strain (SS) of P. xylostella, with synergism ratios that ranged from 1.02 to 1.86. Biochemical studies revealed that the cytochrome P450-dependent monooxygenase increased only 1.13-fold in the metaflu-SEL strain compared with the UNSEL stain; meanwhile, carboxylesterase and glutathione S-transferase activity showed no difference. These results suggest that these detoxification enzymes may be not actively involved in metaflumizone resistance. Furthermore, the metaflu-SEL population showed a moderate level of cross-resistance to indoxacarb (11.63-fold), but only very low cross-resistance to spinosad (1.75-fold), spinetoram (3.52-fold), abamectin (2.81-fold), beta-cypermethrin (0.71-fold), diafenthiuron (0.79-fold), chlorantraniliprole (2.16-fold), BT (WG-001) (3.34-fold), chlorfenapyr (0.49-fold), and chlorfluazuron (0.97-fold). Moreover, metaflumizone resistance decreased from 1087.85- to 1.23-fold in the metaflu-SEL strain after 12 generations without exposure to metaflumizone. These results are useful for formulating insecticide resistance management strategies to control P. xylostella and to delay the development of metaflumizone resistance in the field.
        
Title: Effect of Synergists on Deltamethrin Resistance in the Common Bed Bug (Hemiptera: Cimicidae) Gonzalez-Morales MA, Romero A Ref: J Econ Entomol, 112:786, 2019 : PubMed
The common bed bug, Cimex lectularius L. (Hemiptera: Cimicidae), is an obligate hematophagous insect that has resurged worldwide since the early 2000s. Bed bug control is largely based on the widespread, intensive application of pyrethroid-based insecticide formulations, resulting in the emergence of insecticide-resistant bed bug populations. Insecticide resistance is frequently linked to metabolic detoxification enzymes such as cytochrome monooxygenase (P450s), esterases, glutathione S-tranferase, and carboxylesterase. Therefore, one way to overcome insecticide resistance could be the formulation of insecticides with synergists that counteract metabolic resistance. To test this hypothesis, we evaluated the impact of four synergists-piperonyl butoxide (PBO), diethyl maleate (DEM), S,S,S-tributyl phosphorotrithioate (DEF), and triphenyl phosphate (TPP)-on deltamethrin efficacy in two pyrethroid-resistant bed bug strains. A statistically significant difference in synergism ratios (SR) of a highly resistant field-derived strain (Jersey City, resistance ratio [RR] = 20,000) was noted when any of the four synergists (PBO SR = 20.5; DEM SR = 11.7; DEF SR = 102.3; and TPP SR = 9.7) were used with deltamethrin. In a less deltamethrin-resistant strain, Cincinnati (RR = 3,333), pretreatment with PBO and DEM significantly synergized deltamethrin (PBO SR = 158.8; DEM = 58.8), whereas application of DEF and TPP had no synergistic effect. The synergism data collected strongly suggest that detoxification enzymes play a significant role in the metabolic mechanisms that mediate deltamethrin resistance in bed bugs. The development and use of safe metabolic synergists that suppress detoxification enzymes offers an interesting avenue for the management of insecticide-resistant field populations.
        
Title: Overexpression of multiple cytochrome P450 genes associated with sulfoxaflor resistance in Aphis gossypii Glover Ma K, Tang Q, Zhang B, Liang P, Wang B, Gao X Ref: Pestic Biochem Physiol, 157:204, 2019 : PubMed
Sulfoxaflor is the first commercially available sulfoximine insecticide, which exhibits highly efficacy against many sap-feeding insect pests and has been applied as an alternative insecticide against cotton aphid in China. This study was conducted to investigate the risk of resistance development, the cross-resistance pattern and the potential resistance mechanisms of sulfoxaflor in Aphis gossypii. A colony (SulR strain) of A. gossypii with 245-fold resistance, originated from Xinjiang field population, was established by continuous selection using sulfoxaflor. The SulR strain has developed cross-resistance to imidacloprid (80.8-fold), acetamiprid (19.3-fold), thiamethoxam (10.0-fold), and flupyradifurone (107.5-fold), while no cross-resistance was detected to malathion, omethoate, bifenthrin, methomyl, and carbosulfan. Piperonyl butoxide and S, S, S-tributyl phosphorotrithioate could significantly increase the toxicity of sulfoxaflor to the SulR strain by 5.99- and 4.18-fold, respectively, whereas no synergistic effect with diethyl maleate was observed. The activities of P450s and carboxylesterase were significantly higher in the SulR strain than that in the SS strain. Further gene expression determination demonstrated that nine P450 genes were significantly increased in SulR strain and suppression the expression of CYP6CY13 and CYP6CY19 by RNAi significantly increased the susceptibility of SulR adult aphids to sulfoxaflor. These results demonstrated that the enhancing detoxification by cytochrome P450 monooxygenase may be involved in A.gossypii resistance to sulfoxaflor.
BACKGROUND: Culex mosquitoes cause considerable biting nuisance and sporadic transmission of arboviral and filarial diseases. METHODS: Using standard World Health Organization procedures, insecticide resistance profiles and underlying mechanisms were investigated during dry and wet seasons of 2015 and 2016 in Culex pipiens complex from three neighbouring administrative wards in Ulanga District, Tanzania. Synergist tests with piperonyl butoxide, diethyl maleate, and triphenyl phosphate, were employed to investigate mechanisms of the observed resistance phenotypes. Proportional biting densities of Culex species, relative to other taxa, were determined from indoor surveillance data collected in 2012, 2013, and 2015. RESULTS: Insecticide resistance varied significantly between wards and seasons. For example, female mosquitoes in one ward were susceptible to bendiocarb and fenitrothion in the wet season, but resistant during the dry season, while in neighbouring ward, the mosquitoes were fully susceptible to these pesticides in both seasons. Similar variations occurred against bendiocarb, DDT, deltamethrin, and lambda-cyhalothrin. Surprisingly, with the exception of one ward in the wet season, the Culex populations were susceptible to permethrin, commonly used on bednets in the area. No insecticide resistance was observed against the organophosphates, pirimiphos-methyl and malathion, except for one incident of reduced susceptibility in the dry season. Synergist assays revealed possible involvement of monooxygenases, esterases, and glutathione S-transferase in pyrethroid and DDT resistance. Morphology-based identification and molecular assays of adult Culex revealed that 94% were Cx. pipiens complex, of which 81% were Cx. quinquefasciatus, 2% Cx. pipiens, and 3% hybrids. About 14% of the specimens were non-amplified during molecular identifications. Female adults collected indoors were 100% Cx. pipiens complex, and constituted 79% of the overall biting risk. CONCLUSIONS: The Cx. pipiens complex constituted the greatest biting nuisance inside people's houses, and showed resistance to most public health insecticides possible. Resistance varied at a fine geographical scale, between adjacent wards, and seasons, which warrants some modifications to current insecticide resistance monitoring strategies. Resistance phenotypes are partly mediated by metabolic mechanisms, but require further evaluation through biochemical and molecular techniques. The high densities and resistance in Culex could negatively influence the acceptability of other interventions such as those used against malaria mosquitoes.
Rhipicephalus sanguineus (Latreille) (Ixodida: Ixodidae) is a three-host dog tick found worldwide that is able to complete its' entire lifecycle indoors. Options for the management of R. sanguineus are limited and its' control relies largely on only a few acaricidal active ingredients. Previous studies have confirmed permethrin resistance and fipronil tolerance in R. sanguineus populations, commonly conferred by metabolic detoxification or target site mutations. Herein, five strains of permethrin-resistant and three strains of fipronil-tolerant ticks were evaluated for metabolic resistance using synergists to block metabolic enzymes. Synergist studies were completed with triphenyl phosphate (TPP) for esterase inhibition, piperonyl butoxide (PBO) for cytochrome P450 inhibition, and diethyl maleate (DEM) for glutathione-S-transferase inhibition. Additionally, increased esterase activity was confirmed using gel electrophoresis. The most important metabolic detoxification mechanism in permethrin-resistant ticks was increased esterase activity, followed by increased cytochrome P450 activity. The inhibition of metabolic enzymes did not have a marked impact on fipronil-tolerant tick strains.
        
Title: Pyrethroid resistance in Phytoseiulus macropilis (Acari: Phytoseiidae): cross-resistance, stability and effect of synergists Queiroz MC, Sato ME Ref: Exp Appl Acarol, 68:71, 2016 : PubMed
Phytoseiulus macropilis Banks (Acari: Phytoseiidae) is an effective predator of Tetranychus urticae Koch (Acari: Tetranychidae). The objectives of this research were to study the stability of fenpropathrin resistance and the cross-resistance relationships with different pyrethroids, and also to evaluate the effect of synergists [piperonyl butoxide (PBO), diethyl maleate (DEM) and S,S,S-tributyl phosphorotrithioate (DEF)] on fenpropathrin resistant and susceptible strains of this predaceous mite. The stability of fenpropathrin resistance was studied under laboratory conditions, using P. macropilis populations with initial frequencies of 75 and 50% of resistant mites. The percentages of fenpropathrin resistant mites were evaluated monthly for a period of up to 12 months. A trend toward decreased resistance frequencies was observed only during the first 3-4 months. After this initial decrease, the fenpropathrin resistance was shown to be stable, maintaining constant resistance frequencies (around 30%) until the end of the evaluation period. Toxicity tests carried out using fenpropathrin resistant and susceptible strains of P. macropilis indicated strong positive cross-resistance between fenpropathrin and the pyrethroids bifenthrin and deltamethrin. Bioassays with the synergists DEM, DEF and PBO were also performed. The maximum synergism ratio (SR = LC50 without synergist/LC50 with synergist) detected for the three evaluated synergists (PBO, DEM, DEF) was 5.86 (for DEF), indicating low influence of enzyme detoxification processes in fenpropathrin resistance.
The control of the most important pest of stored maize, the weevil Sitophilus zeamais Motschulsky (Coleoptera: Curculionidae), is mainly achieved with the use of pyrethroid insecticides. However, the intensive use of these compounds has led to the selection of resistant populations and has compromised the control efficacy of this insect pest. Here, the toxicity of indoxacarb for a potential use in the control of S. zeamais was assessed on 13 Brazilian populations. Concentration-mortality bioassays, in the presence of synergists (piperonyl butoxide, triphenyl phosphate, and diethyl maleate), were used to assess potential metabolic-based indoxacarb resistance mechanisms. We also assessed the behavioral (locomotory) responses of these populations to indoxacarb exposure. The results showed significant differences between the populations (LD50 values ranged from 0.06 to 13.99 mg a.i/kg of grains), resulting in resistance ratios of >200-fold between the least (Canarana-MT) and the most (Espirito Santo do Pinhal-SP) susceptible populations. The results obtained with synergized indoxacarb suggest the involvement of esterases and glutathione-S-transferases on indoxacarb action, and also suggest the involvement of cytochrome P450-dependent monooxygenases as a potential indoxacarb resistance mechanism in Brazilian populations of S. zeamais. Although indoxacarb-induced behavioral avoidance varied among populations, some resistant populations (e.g., Canarana-MT) were able to reduce exposure to indoxacarb by spending more time in the nontreated areas. Collectively, our findings indicate that the behavioral (locomotory) and physiological responses of these insects may compromise the control efficacy of oxadiazine insecticides (e.g., indoxacarb) in Brazilian populations of S. zeamais.
        
Title: Insecticides resistance in the Culex quinquefasciatus populations from northern Thailand and possible resistance mechanisms Yanola J, Chamnanya S, Lumjuan N, Somboon P Ref: Acta Trop, 149:232, 2015 : PubMed
The mosquito vector Culex quinquefasciatus is known to be resistant to insecticides worldwide, including Thailand. This study was the first investigation of the insecticide resistance mechanisms, involving metabolic detoxification and target site insensitivity in C. quinquefasciatus from Thailand. Adult females reared from field-caught larvae from six provinces of northern Thailand were determined for resistant status by exposing to 0.05% deltamethrin, 0.75% permethrin and 5% malathion papers using the standard WHO susceptibility test. The overall mortality rates were 45.8%, 11.4% and 80.2%, respectively. A fragment of voltage-gated sodium channel gene was amplified and sequenced to identify the knock down resistance (kdr) mutation. The ace-1 gene mutation was determined by using PCR-RFLP. The L1014F kdr mutation was observed in all populations, but the homozygous mutant F/F1014 genotype was found only in two of the six provinces where the kdr mutation was significantly correlated with deltamethrin resistance. However, none of mosquitoes had the G119S mutation in the ace-1 gene. A laboratory deltamethrin resistant strain, Cq_CM_R, has been established showing a highly resistant level after selection for a few generations. The mutant F1014 allele frequency was significantly increased after one generation of selection. A synergist assay was performed to assess the metabolic detoxifying enzymes. Addition of bis(4-nitrophenyl)-phosphate (BNPP) and diethyl maleate (DEM), inhibitors of esterases and glutathione S-transferases (GST), respectively, into the larval bioassay of the Cq_CM strain with deltamethrin showed no significant reduction. By contrast, addition of piperonyl butoxide (PBO), an inhibitor of cytochrome P450 monooxygenases, showed a 9-fold reduction of resistance. Resistance to pyrethroids in C. quinquefasciatus is widely distributed in northern Thailand. This study reports for the first time for the detection of the L1014F kdr mutation in wild populations of C. quinquefasciatus in Thailand. At least two major mechanisms, kdr and cytochrome P450 monooxygenases, confer resistance to deltamethrin in Thai C. quinquefasciatus populations.
        
Title: Thiamethoxam resistance selected in the western flower thrips Frankliniella occidentalis (Thysanoptera: Thripidae): Cross-resistance patterns, possible biochemical mechanisms and fitness costs analysis Gao CF, Ma SZ, Shan CH, Wu SF Ref: Pestic Biochem Physiol, 114:90, 2014 : PubMed
The western flower thrips (WFT) Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), an important pest of various crops in the world, has invaded China since 2003. To understand the risks and to determine possible mechanisms of resistance to thiamethoxam in WFT, a resistant strain was selected under the laboratory conditions. Cross-resistance and the possible biochemical resistance mechanisms were investigated in this study. A 15.1-fold thiamethoxam-resistant WFT strain (TH-R) was established after selection for 55 generations. Compared with the susceptible strain (TH-S), the selected TH-R strain showed extremely high level cross-resistance to imidaclothiz (392.1-fold) and low level cross-resistance to dinotefuran (5.7-fold), acetamiprid (2.9-fold) and emamectin benzoate (2.1-fold), respectively. No cross-resistance to other fourteen insecticides was detected. Synergism tests showed that piperonyl butoxide (PBO) and triphenyl phosphate (TPP) produced a high synergism of thiamethoxam effects in the TH-R strain (2.6- and 2.6-fold respectively). However, diethyl maleate (DEM) did not act synergistically with thiamethoxam. Biochemical assays showed that mixed function oxidase (MFO) activities and carboxylesterase (CarE) activity of the TH-R strain were 2.8- and 1.5-fold higher than that of the TH-S strain, respectively. When compared with the TH-S strain, the TH-R strain had a relative fitness of 0.64. The results show that WFT develops resistance to thiamethoxam after continuous application and thiamethoxam resistance had considerable fitness costs in the WFT. It appears that enhanced metabolism mediated by cytochrome P450 monooxygenases and CarE was a major mechanism for thiamethoxam resistance in the WFT. The use of cross-resistance insecticides, including imidaclothiz and dinotefuran, should be avoided for sustainable resistance management.
        
Title: Enzymes mediating resistance to lambda-cyhalothrin in Eriopis connexa (Coleoptera: Coccinellidae) Rodrigues AR, Siqueira HA, Torres JB Ref: Pestic Biochem Physiol, 110:36, 2014 : PubMed
Resistance to widely used insecticide, lambda-cyhalothrin, was recently reported in the predatory lady beetle Eriopis connexa (Germar) (Coleoptera: Coccinellidae). However, to understand whether metabolic mechanisms underlie such resistance, synergism bioassays and in vitro studies were carried out by using inhibitors and model substrates for enzymatic assays, respectively. The LD50s estimated for susceptible and resistant populations etag of lambda-cyhalothrin/insect, and thus, a 22-fold difference in resistance ratio. Synergism ratios for the susceptible population with piperonyl butoxide (PBO), diethyl maleate (DEM), triphenyl phosphate (TPP), and S,S,S-tributylphosphorotrithioate (DEF) were respectively 33.8-, 0.24-, 0.35-, and 4.25-fold, while for the resistant population, they were 1463.0-, 0.79-, 0.85-, and 282.6-fold, respectively. The synergized resistance ratios were 0.50-, 2.00-, 6.75-, and 8.77-fold with PBO, DEF, DEM, and TPP, respectively, while resistance was virtually suppressed with DEF. The esterase exhibited 4.16-, 4.03-, and 5.38-fold greater activity towards formation of alpha-naphthol, beta-naphthol, and 4-nitrophenol in the resistant population of E. connexa than in the susceptible population. The activity of esterase depended on concentrations of DEF applied, either using alpha-naphthol or beta-naphthol, which completely inhibited the activity at 636etaM. The PBO inhibited the beta-naphthol formation in approximately 50%, suggesting it as inhibitor of esterases. The activities of glutathione-S-transferase were similar and corresponded to 0.36-0.47etamol(-1)min(-1)mug of protein, for S and R populations, respectively. Similarly, the activities of cytochrome P450-dependent microsomal monooxygenases were 0.04 and 0.05etamol(-1)min(-1)mug of protein. The native gel indicated that the formation of beta-naphthol was completely inhibited by methyl-paraoxon, but only partially inhibited by eserine, TPP, and PBO. Although other studies with DEF and PBO have demonstrated strong inhibition of type B carboxylesterase associated with insecticide resistance, the results reported here do not rule out metabolism by cytochrome P450-dependent microsomal monooxygenases as a factor conferring E. connexa resistance to lambda-cyhalothrin and confirmed that PBO may also act by inhibiting esterases of insects.
        
Title: [Resistance mechanisms and cross-resistance of phoxim-resistant Frankliniella occidentalis Pergande population] Wang SY, Zhou XH, Zhang AS, Li LL, Men XY, Zhang SC, Liu YJ, Yu Y Ref: Ying Yong Sheng Tai Xue Bao, 23:1933, 2012 : PubMed
To understand the resistance risks of Frankliniella occidentalis Pergande against phoxim, this paper studied the resistance mechanisms of phoxim-resistant F. occidentalis population against phoxim and the cross-resistance of the population against other insecticides. The phoxim-resistant population had medium level cross-resistance to chlorpyrifos, lambda-cyhalothrin, and methomyl, low level cross-resistance to chlorfenapyr, imidacloprid, emamectin-benzoate, and spinosad, but no cross-resistance to acetamiprid and abamectin. The synergists piperonyl butoxide (PBO), s, s, s-tributyl phosphorotrithioate (DEF), and triphenyl phosphate (TPP) had significant synergism (P < 0.05) on the toxicity of phoxim to the resistant (XK), field (BJ), and susceptible (S) populations, while diethyl maleate (DEM) had no significant synergism to XK and S populations but had significant synergism to BJ population. As compared with S population, the XK and BJ populations had significantly increased activities of mixed-functional oxidases P450 (2.79-fold and 1.48-fold), b, (2.88-fold and 1.88-fold), O-demethylase (2.60-fold and 1.68-fold), and carboxylesterase (2.02-fold and 1.61-fold, respectively), and XK population had a significantly increased acetylcholine esterase activity (3.10-fold). Both XK and BJ population had an increased activity of glutathione S-transferases (1.11-fold and 1.20-fold, respectively), but the increment was not significant. The increased detoxification enzymes activities in F. occidentalis could play an important role in the resistance of the plant against phoxim.
        
Title: Cross-resistance of bisultap resistant strain of Nilaparvata lugens and its biochemical mechanism Ling S, Zhang R Ref: J Econ Entomol, 104:243, 2011 : PubMed
The resistant (R) strain of the planthopper Nilaparvata lugens (Stal) selected for bisultap resistance displayed 7.7-fold resistance to bisultap and also had cross-resistance to nereistoxin (monosultap, thiocyclam, and cartap), chlorpyrifos, dimethoate, and malathion but no cross-resistance to buprofezin, imidacloprid, and fipronil. To find out the biochemical mechanism of resistance to bisultap, biochemical assay was done. The results showed that cytochrome P450 monooxygenases (P450) activity in R strain was 2.71-fold that in susceptible strain (S strain), in which the changed activity for general esterase (EST) was 1.91 and for glutathione S-transferases only 1.32. Piperonyl butoxide (PBO) could significantly inhibit P450 activity (percentage of inhibition [PI]: 37.31%) in the R strain, with ESTs PI = 16.04% by triphenyl phosphate (TPP). The results also demonstrated that diethyl maleate had no synergism with bisultap. However, PBO displayed significant synergism in three different strains, and the synergism increased with resistance (S strain 1.42, Lab strain, 2.24 and R strain, 3.23). TPP also showed synergism for three strains, especially in R strain (synergistic ratio = 2.47). An in vitro biochemical study and in vivo synergistic study indicated that P450 might be play important role in the biochemical mechanism of bisultap resistance and that esterase might be the important factor of bisultap resistance. Acetylcholinesterase (AChE) insensitivity play important role in bisultap resistance. We suggest that buprofezin, imidacloprid, and fipronil could be used in resistance management programs for N. lugens via alternation and rotation with bisultap.
        
Title: Cross-resistance study and biochemical mechanisms of thiamethoxam resistance in B-biotype Bemisia tabaci (Hemiptera: Aleyrodidae) Feng Y, Wu Q, Wang S, Chang X, Xie W, Xu B, Zhang Y Ref: Pest Manag Sci, 66:313, 2010 : PubMed
BACKGROUND: B-biotype Bemisia tabaci (Gennadius) has invaded China over the past two decades. To understand the risks and to determine possible mechanisms of resistance to thiamethoxam in B. tabaci, a resistant strain was selected in the laboratory. Cross-resistance and the biochemical mechanisms of thiamethoxam resistance were investigated in the present study. RESULTS: A 66.3-fold thiamethoxam-resistant B. tabaci strain (TH-R) was established after selection for 36 generations. Compared with the susceptible strain (TH-S), the selected TH-R strain showed obvious cross-resistance to imidacloprid (47.3-fold), acetamiprid (35.8-fold), nitenpyram (9.99-fold), abamectin (5.33-fold) and carbosulfan (4.43-fold). No cross-resistance to fipronil, chlorpyrifos or deltamethrin was seen. Piperonyl butoxide (PBO) and triphenyl phosphate (TPP) exhibited significant synergism on thiamethoxam effects in the TH-R strain (3.14- and 2.37-fold respectively). However, diethyl maleate (DEM) did not act synergistically with thiamethoxam. Biochemical assays showed that cytochrome P450 monooxygenase activities increased 1.21- and 1.68-fold respectively, and carboxylesterase activity increased 2.96-fold in the TH-R strain. However, no difference was observed for glutathione S-transferase between the two strains. CONCLUSION: B-biotype B. tabaci develops resistance to thiamethoxam. Cytochrome P450 monooxygenase and carboxylesterase appear to be responsible for the resistance. Reasonable resistance management that avoids the use of cross-resistance insecticides may delay the development of resistance to thiamethoxam in this species.
        
Title: Resistance selection and biochemical mechanism of resistance to two Acaricides in Tetranychus cinnabarinus (Boiduval) Lin H, Chuan-hua X, Jin-jun W, Ming L, Wen-cai L, Zhi-mo Z Ref: Pesticide Biochemistry and Physiology, 93:47, 2009 : PubMed
A Tetranychus cinnabarinus strain was collected from Chongqing, China. After 42 generations of selection with abamectin and 20 generations of selection with fenpropathrin in the laboratory, this T. cinnabarinus strain developed 8.7- and 28.7-fold resistance, respectively. Resistance to abamectin in AbR (abamectin resistant strain) and to fenpropathrin in FeR (fenpropathrin resistant strain) was partially suppressed by piperonyl butoxide (PBO), diethyl maleate (DEM) and triphenyl phosphate (TPP), inhibitors of mixed function oxidase (MFO), glutathione S-transferases (GST), and hydrolases, respectively, suggesting that these three enzyme families are important in conferring abamectin and fenpropathrin resistance in T. cinnabarinus. The major resistant mechanism to abamectin was the increasing activities of carboxylesterases (CarE), glutathione-S-transferase (GST) and mixed function oxidase (MFO), and the activity in resistant strain developed 2.7-, 3.4- and 1.4-fold contrasted to that in susceptible strain, respectively. The activity of glutathione-S-transferase (GST) in the FeR strain developed 2.8-fold when compared with the susceptible strain, which meant the resistance to fenpropathrin was related with the activity increase of glutathione-S-transferase (GST) in T. cinnabarinus. The result of the kinetic mensuration of carboxylesterases (CarE) showed that the structure of CarE in the AbR has been changed.
        
Title: Effects of spinosad on Helicoverpa armigera (Lepidoptera: Noctuidae) from China: tolerance status, synergism and enzymatic responses Wang D, Qiu X, Ren X, Zhang W, Wang K Ref: Pest Manag Sci, 65:1040, 2009 : PubMed
BACKGROUND:
Spinosad is increasingly used in pest management programmes, and resistance to it has been detected in recent years. However, there is no report on the susceptibilities of field populations of Helicoverpa armigera (Hbner) from China. Furthermore, the impact of spinosad on metabolic enzymes in this pest remains unknown.
RESULTS:
Four populations of H. armigera from different locations in China displayed less than 6.5-fold difference in LC(50) to spinosad, the highest being in the Xinjiang population, followed by Xiajin, Taian and Hubei populations, while there was no significant difference at LC(99) level among the four populations. The toxicity of spinosad could be synergised by piperonyl butoxide (PBO) and triphenylphosphate (TPP), but not by diethyl maleate (DEM). Spinosad exposure for 48 h significantly increased the activities of p-nitroanisole O-demethylase (ODM), while no significant changes in glutathione-S-transferase (GST) and carboxyl esterase (CarE) were observed.
CONCLUSION:
Field populations of H. armigera from China displayed marginally different susceptibilities to spinosad and had a relatively low LC(50). Cytochrome P450 monooxygenase might be involved in the metabolism of, and hence resistance to, spinosad in this pest in China.
        
Title: Mechanisms of organophosphate resistance in a field population of oriental migratory locust, Locusta migratoria manilensis (Meyen) Yang ML, Zhang JZ, Zhu KY, Xuan T, Liu XJ, Guo YP, Ma EB Ref: Archives of Insect Biochemistry & Physiology, 71:3, 2009 : PubMed
The susceptibilities to three organophosphate (OP) insecticides (malathion, chlorpyrifos, and phoxim), responses to three metabolic synergists [triphenyl phosphate (TPP), piperonyl butoxide (PBO), and diethyl maleate (DEM)], activities of major detoxification enzymes [general esterases (ESTs), glutathione S-transferases (GSTs), and cytochrome P450 monooxygenases (P450s)], and sensitivity of the target enzyme acetylcholinesterase (AChE) were compared between a laboratory-susceptible strain (LS) and a field-resistant population (FR) of the oriental migratory locust, Locusta migratoria manilensis (Meyen). The FR was significantly resistant to malathion (57.5-fold), but marginally resistant to chlorpyrifos (5.4) and phoxim (2.9). The malathion resistance of the FR was significantly diminished by TPP (synergism ratio: 16.2) and DEM (3.3), but was unchanged by PBO. In contrast, none of these synergists significantly affected the toxicity of malathion in the LS. Biochemical studies indicated that EST and GST activities in the FR were 2.1- to 3.2-fold and 1.2- to 2.0-fold, respectively, higher than those in the LS, but there was no significant difference in P450 activity between the LS and FR. Furthermore, AChE from the FR showed 4.0-fold higher activity but was 3.2-, 2.2-, and 1.1-fold less sensitive to inhibition by malaoxon, chlorpyrifos-oxon, and phoxim, respectively, than that from the LS. All these results clearly indicated that the observed malathion resistance in the FR was conferred by multiple mechanisms, including increased detoxification by ESTs and GSTs, and increased activity and reduced sensitivity of AChE to OP inhibition.
Inhibition of bovine erythrocyte acetylcholinesterase (free and immobilized on controlled pore glass) by separate and simultaneous exposure to malathion and malathion transformation products which are generally formed during storage or through natural or photochemical degradation was investigated. Increasing concentrations of malathion, its oxidation product malaoxon, and its isomerisation product isomalathion inhibited free and immobilized AChE in a concentration-dependent manner. KI, the dissociation constant for the initial reversible enzyme inhibitor-complex, and k3, the first order rate constant for the conversion of the reversible complex into the irreversibly inhibited enzyme, were determined from the progressive development of inhibition produced by reaction of native AChE with malathion, malaoxon and isomalathion. KI values of 1.3 x 10(-4) M(-1), 5.6 x 10(-6) M(-1) and 7.2 x 10(-6)M(-1) were obtained for malathion, malaoxon and isomalathion, respectively. The IC50 values for free/immobilized AChE, (3.7 +/- 0.2) x 10(-4) M/(1.6 +/-0.1) x 10(-4), (2.4 +/- 0.3) x 10(-6)/(3.4 +/- 0.1) x 10(-6)M and (3.2 +/- 0.3) x 10(-6) M/(2.7 +/- 0.2) x 10(-6) M, were obtained from the inhibition curves induced by malathion, malaoxon and isomalathion, respectively. However, the products formed due to photoinduced degradation, phosphorodithioic O,O,S-trimethyl ester and O,O-dimethyl thiophosphate, did not noticeably affect enzymatic activity, while diethyl maleate inhibited AChE activity at concentrations > 10mM. Inhibition of acetylcholinesterase increased with the time of exposure to malathion and its inhibiting by-products within the interval from 0 to 5 minutes. Through simultaneous exposure of the enzyme to malaoxon and isomalathion, an additive effect was achieved for lower concentrations of the inhibitors (in the presence of malaoxon/isomalathion at concentrations 2 x 10(-7) M/2 x 10(-7) M, 2 x 10(-7) M/3 x 10(-7)M and 2 x 10(-7) M/4.5 x 109-7) M), while an antagonistic effect was obtained for all higher concentrations of inhibitors. The presence of a non-inhibitory degradation product (phosphorodithioic O,O,S-trimethyl ester) did not affect the inhibition efficiencies of the malathion by-products, malaoxon and isomalathion.
        
Title: Inhibition of AChE by single and simultaneous exposure to malathion and its degradation products Krstic D, Colovic M, Krinulovic K, Djuric D, Vasic V Ref: Gen Physiol Biophys, 26:247, 2007 : PubMed
In vitro inhibition of bovine erythrocytes acetylcholinesterase (AchE) by separate and simultaneous exposure to organophosphorous insecticide malathion and the transformation products, which are generally formed during the storage or natural as well as photochemical degradation pathways of malathion, was investigated. The increasing concentration of malathion, its oxidation product - malaoxon and isomerisation product - isomalathion inhibited AChE activity in a concentration-dependent manner. The half-maximum inhibitory concentrations (IC(50) values): (3.2 +/- 0.1) x 10(-5) mol/l, (4.7 +/- 0.8) x 10(-7) mol/l and (6.0 +/- 0.5) x 10(-7)mol/l were obtained from the inhibition curves induced by malathion, malaoxon and isomalathion, respectively. However, the products formed due to photoinduced degradation, phosphorodithioic O,O,S-trimethyl phosphorodithioic ester (OOS(S)) and O,O-dimethyl thiophosphate did not noticeably affect the enzyme activity at all investigated concentrations, while diethyl maleate inhibited the AChE activity at concentrations >10 mmol/l. By simultaneous exposure of the enzyme to malaoxon and isomalathion in various concentration combinations the additive effect was achieved by low concentration of inhibitors, while the antagonistic effect was obtained at high concentration (>or= 3 x 10(-7) mol/l) of inhibitors. Inhibitory power of irradiated samples of 1 +/- 10(-5) mol/l malathion can be attributed to the formation of malaoxon and isomalathion, organophosphates about 100 times more toxic than their parent compound, while the presence of non-inhibiting degradation product OOS(S) did not affect the inhibitor efficiency of inhibiting malathion by-products, malaoxon and isomalathion.
        
Title: Insecticide toxicity and synergism by enzyme inhibitors in 18 species of pest insect and natural enemies in crucifer vegetable crops Wu G, Miyata T, Kang CY, Xie LH Ref: Pest Manag Sci, 63:500, 2007 : PubMed
The toxicities of three enzyme inhibitors and their synergistic effects on four insecticides were studied by using the dry film method on field populations of 18 species of insects collected in Jianxin and Shanjie, China, from 2003 to 2005. Meanwhile, the inhibitory effects of these enzyme inhibitors on the activities of acetylcholinesterases (AChE), carboxyesterases (CarE) and glutathione-S-transferases (GST), in vivo, were also studied. In general, triphenyl phosphate (TPP) and diethyl maleate (DEM) showed low toxicities to six herbivorous pest insects, four ladybirds and eight parasitoids. Piperonyl butoxide (PB) exhibited low toxicities to the herbivorous pest insects and ladybirds, but high toxicities to the eight parasitoids. The tolerance to the insecticides in 11 pest insects and natural enemies was mainly associated with the tolerance to PB. PB showed the highest synergism on methamidophos, fenvalerate, fipronil and avermectin in nine species of pest insects and natural enemies. In general, TPP and DEM showed significant synergisms to these four insecticides in four parasitoid species. However, in contrast to their effects on the parasitoids, the synergistic effects of TPP and DEM on the four insecticides by TPP and DEM against four pest insects and one ladybird varied depending on the insect species and enzyme inhibitor. Activity of AChE, CarE or GST could be strongly inhibited, in vivo, by PB, TPP or DEM, depending on the insect species and enzyme inhibitors. From the results obtained in this study, mixed-function oxidase (MFO) was thought to play the most critical role in insect tolerances to the tested insecticides in the field. Low competition existed in the evolution of insecticide resistance in the field populations of parasitoids, as compared with herbivorous pest insects and ladybirds. Possible causes of the high synergistic effects of PB on the four classes of insecticides, based on multiattack on the activity of CarE, GST or AChE in the insect species, are also discussed.
        
Title: Mediation of pyrethroid insecticide toxicity to honey bees (Hymenoptera: Apidae) by cytochrome P450 monooxygenases Johnson RM, Wen Z, Schuler MA, Berenbaum MR Ref: J Econ Entomol, 99:1046, 2006 : PubMed
Honey bees, Apis mellifera L., often thought to be extremely susceptible to insecticides in general, exhibit considerable variation in tolerance to pyrethroid insecticides. Although some pyrethroids, such as cyfluthrin and lambda-cyhalothrin, are highly toxic to honey bees, the toxicity of tau-fluvalinate is low enough to warrant its use to control parasitic mites inside honey bee colonies. Metabolic insecticide resistance in other insects is mediated by three major groups of detoxifying enzymes: the cytochrome P450 monooxygenases (P450s), the carboxylesterases (COEs), and the glutathione S-transferases (GSTs). To test the role of metabolic detoxification in mediating the relatively low toxicity of tau-fluvalinate compared with more toxic pyrethroid insecticides, we examined the effects of piperonyl butoxide (PBO), S,S,S-tributylphosphorotrithioate (DEF), and diethyl maleate (DEM) on the toxicity of these pyrethroids. The toxicity of the three pyrethroids to bees was greatly synergized by the P450 inhibitor PBO and synergized at low levels by the carboxylesterase inhibitor DEF. Little synergism was observed with DEM. These results suggest that metabolic detoxification, especially that mediated by P450s, contributes significantly to honey bee tolerance of pyrethroid insecticides. The potent synergism between tau-fluvalinate and PBO suggests that P450s are especially important in the detoxification of this pyrethroid and explains the ability of honey bees to tolerate its presence.
        
Title: Parasitization by Cotesia plutellae enhances detoxifying enzyme activity in Plutella xylostella Takeda T, Nakamatsu Y, Tanaka T Ref: Pesticide Biochemistry and Physiology, 86:15, 2006 : PubMed
Insecticidal tests using diazinon showed that the mortality of Plutella xylostella larvae parasitized by Cotesia plutellae was reduced by 4.6-fold compared to that of the nonparasitized hosts. The use of chemicals with synergistic effect to insecticides in toxicity assay helps to elucidate the kind of enzyme involved in lowering insect mortality. Synergism of diethyl maleate and piperonyl butoxide with diazinon resulted to 2.4- and 1.9-fold increase, respectively, in susceptibility of parasitized larvae compared to those of nonparasitized larvae. These results indicated the possibility that the decrease in susceptibility to diazinon was due to the elevated activities of glutathione-S-transferase (GST) and cytochrome P450 monooxygenase (CYP), respectively. The GST activities in parasitized larvae were significantly higher than those of nonparasitized ones starting from three days post-parasitization until emergence of parasitoid larva. High GST activities during late parasitism could be attributed to both enzyme activities toward diazinon of parasitized P. xylostella larva itself and C. plutellae larva inside larval host. High GST activity one day after parasitization, although statistical significance was not detected, was caused by polydnavirus (PDV) and the venom of C. plutellae not by parasitoid larvae. Artificial injection of PDV plus venom demonstrated that the resulting increase in GST activity is similar to the increase brought by parasitization. High CYP activity after 3 days post-parasitization in parasitized larva was attributed mainly to the activity of parasitoid larva. Carboxylesterase activity in the parasitized host remained at a high level, while that in the nonparasitized host decreased slightly as pupation approaches. On the other hand, acetylcholinesterase activity also remained constant after parasitization until larval emergence, while that of the nonparasitized hosts decreased gradually as the host larvae approach pupation. These results were supported by inhibition tests using diazoxon in vitro.
The interactions between six insecticides (methiocarb, formetanate, acrinathrin, deltamethrin, methamidophos and endosulfan) and three potential synergists (piperonyl butoxide (PBO), S,S,S-tributyl phosphorotrithioate (DEF) and diethyl maleate (DEM)) were studied by topical exposure in strains selected for resistance to each insecticide, and in a susceptible strain of Frankliniella occidentalis (Pergande). In the susceptible strain PBO produced appreciable synergism only of formetanate, methiocarb and methamidophos. Except for endosulfan, PBO synergized all the insecticides to varying degrees in the resistant strains. A very high level of synergism by PBO was found with acrinathrin, which reduced the resistance level from 3344- to 36-fold. PBO slightly synergized the carbamates formetanate (4.6-fold) and methiocarb (3.3-fold). PBO also produced a high synergism of deltamethrin (12.5-fold) and methamidophos (14.3-fold) and completely restored susceptibility to both insecticides. DEF did not produce synergism with any insecticide in the resistant strains and DEM was slightly synergistic to endosulfan (3-fold). These studies indicate that an enhanced detoxification, mediated by cytochrome P-450 monooxygenases, is the major mechanism imparting resistance to different insecticides in F occidentalis. Implications of different mechanisms in insecticide resistance in F occidentalis are discussed.
        
Title: Chlorpyrifos resistance in mosquito Culex quinquefasciatus Liu H, Xu Q, Zhang L, Liu N Ref: Journal of Medical Entomology, 42:815, 2005 : PubMed
Two mosquito strains of Culex quinquefasciatus Say, MAmCq and HAmCq, were collected from Mobile and Huntsville, AL, respectively, after the control of mosquitoes with insecticides proved difficult. A synergism study showed that resistance to chlorpyrifos in MAmCq and HAmCq was not suppressed by piperonyl butoxide (PBO) and S,S,S,-tributylphosphorotrithioate (DEF), suggesting that P450 monooxygenase- and hydrolase-mediated detoxication does not contribute to chlorpyrifos resistance in either strain. Diethyl maleate (DEM) did not cause any significant change in the level of chlorpyrifos toxicity to HAmCq. However, DEM enhanced toxicity of chlorpyrifos to MAmCq 2.5-fold, indicating that glutathione S-transferase (GST)-mediated detoxication may play a minor role in the resistance of MAmCq. An inhibition study of acetylcholinesterase (AChE) by chlorpyrifos showed that bimolecular rate constants (Ki) of chlorpyrifos for the inhibition of AChE in adults and larvae of the susceptible S-Lab strain were 2.2- and 1.9-fold higher, respectively, than in the HAmCq strain and 3.4- and 3.8-fold higher than in the MAmCq strain. The single mutation, G119S, resulting from a single nucleotide polymorphism (SNP), G to A, in ace-1 acetylcholinesterase gene was present in HAmCq and MAmCq mosquitoes. The frequency of the heterozygote for the G119S mutant allele in the HAmCq and MAmCq mosquito populations was 0.25 and 0.45, respectively, and no individuals in either of these mosquito strains were homozygous for the A allele. It thus seems likely that the presence of heterozygous individuals for the G119S allele in HAmCq and MAmCq populations may be a response to the insensitivity of AChE observed in these two mosquito strains.
        
Title: Susceptibilities to methamidophos and enzymatic characteristics in 18 species of pest insects and their natural enemies in crucifer vegetable crops Wu G, Miyata T Ref: Pesticide Biochemistry and Physiology, 82:79, 2005 : PubMed
The susceptibilities to methamidophos as well as the kinetic and inhibitory parameter of acetylcholinesterases (AChE) and the activities of carboxyestsrases (CarE) and glutathione-S-transferases (GST) were studied in 18 species field populations of insects collected in Fuzhou, China during April and May 2000 and 2001. The insect species included five hymenopteran endoparasitoids, one hymenopteran exoparasitoid, one hymenopteran hyperparasitoid, one dipteran predator, four coleopteran predator ladybirds, six herbivorous pest insects of lepidoptera, diptera, homoptera, and coleoptera, respectively. There existed significant correlations between the susceptibility to methamidophos and the ki values of AChE to methamidophos, dichlorvos, and carbofuran and between the ki and Vmax values of AChE among 18 species of insects. The six herbivorous pests and four ladybirds showed significantly low ki and Vmax values of AChE compared to the seven parasitoids and predator Epistrophe balteate. It was difficult to correlate the susceptibility to methamidophos or the ki values with the Km values of AChE, or with the activity of CarE and GST. The activities of CarE and GST varied depending on the different insect species. Significant synergisms of piperonyl butoxide (PB), triphenyl phosphate (TPP), and diethyl maleate (DEM) with methamidophos were observed in 14 pest insects and their natural enemies. Synergisms of PB were found to be the greatest. Reduced ki values suggested that insensitive AChE might play a critical role in the tolerance to methamidophos in the 18 insect species. The detoxification enzymes, mixed-function oxidase (MFO), CarE, and GST, were believed to be involved in the tolerance to methamidophos. MFO might play the most important role, and CarE or GST might be important in the tolerance in some insect species. Different models of tolerance to methamidophos and enzymatic potential were existed in parasitoids, predators, and herbivores based on the different selection of insecticide pressure (either directly by exposing to the spray in the field, or indirectly by the insecticides penetrated into the body of host insects) as well as different ecological and biological habitats.
The transcription factor NF-E2-related factor 2 (Nrf2) plays an essential role in the mammalian response to chemical and oxidative stress through induction of hepatic phase II detoxification enzymes and regulation of glutathione (GSH). Enhanced liver damage in Nrf2-deficient mice treated with acetaminophen suggests a critical role for Nrf2; however, direct evidence for Nrf2 activation following acetaminophen exposure was previously lacking. We show that acetaminophen can initiate nuclear translocation of Nrf2 in vivo, with maximum levels reached after 1 hour, in a dose dependent manner, at doses below those causing overt liver damage. Furthermore, Nrf2 was shown to be functionally active, as assessed by the induction of epoxide hydrolase, heme oxygenase-1, and glutamate cysteine ligase gene expression. Increased nuclear Nrf2 was found to be associated with depletion of hepatic GSH. Activation of Nrf2 is considered to involve dissociation from a cytoplasmic inhibitor, Kelch-like ECH-associated protein 1 (Keap1), through a redox-sensitive mechanism involving either GSH depletion or direct chemical interaction through Michael addition. To investigate acetaminophen-induced Nrf2 activation we compared the actions of 2 other GSH depleters, diethyl maleate (DEM) and buthionine sulphoximine (BSO), only 1 of which (DEM) can function as a Michael acceptor. For each compound, greater than 60% depletion of GSH was achieved; however, in the case of BSO, this depletion did not cause nuclear translocation of Nrf2. In conclusion, GSH depletion alone is insufficient for Nrf2 activation: a more direct interaction is required, possibly involving chemical modification of Nrf2 or Keap1, which is facilitated by the prior loss of GSH.
        
Title: Resistance to coumaphos and diazinon in Boophilus microplus (Acari: Ixodidae) and evidence for the involvement of an oxidative detoxification mechanism Li AY, Davey RB, Miller RJ, George JE Ref: Journal of Medical Entomology, 40:482, 2003 : PubMed
The levels of resistance to two organophosphate acaricides, coumaphos and diazinon, in several Mexican strains of Boophilus microplus (Canestrini) were evaluated using the FAO larval packet test. Regression analysis of LC50 data revealed a significant cross-resistance pattern between those two acaricides. Metabolic mechanisms of resistance were investigated with synergist bioassays. Piperonyl butoxide (PBO) reduced coumaphos toxicity in susceptible strains, but synergized coumaphos toxicity in resistant strains. There was a significant correlation between PBO synergism ratios and the coumaphos resistance ratios. The results suggest that an enhanced cytochrome P450 monooxygenase (cytP450)-mediated detoxification mechanism may exist in the resistant strains, in addition to the cytP450-mediated metabolic pathway that activates coumaphos. PBO failed to synergize diazinon toxicity in resistant strains, suggesting the cytP450 involved in detoxification were specific. Triphenylphosphate (TPP) synergized toxicity of both acaricides in both susceptible and resistant strains, and there was no correlation between TPP synergism ratios and the LC50 estimates for either acaricide. Esterases may not play a major role in resistance to coumaphos and diazinon in those strains. Bioassays with diethyl maleate (DEM) revealed a significant correlation between DEM synergism ratios and LC50 estimates for diazinon, suggesting a possible role for glutathione S-transferases in diazinon detoxification. Resistance to coumaphos in the Mexican strains of B. microplus was likely to be conferred by both a cytP450-mediated detoxification mechanism described here and the mechanism of insensitive acetylcholinesterases reported elsewhere. The results of this study also underscore the potential risk of coumaphos resistance in B. microplus from Mexico to the U.S. cattle fever tick eradication program.
        
Title: Insecticide use and organophosphate resistance in the coffee leaf miner Leucoptera coffeella (Lepidoptera: Lyonetiidae) Fragoso DB, Guedes RN, Picanco MC, Zambolim L Ref: Bull Entomol Res, 92:203, 2002 : PubMed
Increasing rates of insecticide use against the coffee leaf minerLeucoptera coffeella(Guerin-Meneville) and field reports on insecticide resistance led to an investigation of the possible occurrence of resistance of this species to some of the oldest insecticides used against it in Brazil: chlorpyrifos, disulfoton, ethion and methyl parathion. Insect populations were collected from ten sites in the state of Minas Gerais, Brazil and these populations were subjected to discriminating concentrations established from insecticide LC99s estimated for a susceptible standard population. Eight of the field-collected populations showed resistance to disulfoton, five showed resistance to ethion, four showed resistance to methyl parathion, and one showed resistance to chlorpyrifos. The frequency of resistant individuals in each population ranged from 10 to 93% for disulfoton, 53 to 75% for ethion, 23 to 76% for methyl parathion, and the frequency of resistant individuals in the chlorpyrifos resistant population was 35%. A higher frequency of individuals resistant to chlorpyrifos, disulfoton and ethion was associated with greater use of insecticides, especially other organophosphates. This finding suggests that cross-selection, mainly between organophosphates, played a major role in the evolution of insecticide resistance in Brazilian populations of L. coffeella. Results from insecticide bioassays with synergists (diethyl maleate, piperonyl butoxide and triphenyl phosphate) suggested that cytochrome P450-dependent monooxygenases may play a major role in resistance with minor involvement of esterases and glutathione S-transferases.
        
Title: Evaluation of the developmental toxicity of thalidomide using frog embryo teratogenesis assay-xenopus (FETAX): biotransformation and detoxification Fort DJ, Stover EL, Bantle JA, Finch RA Ref: Teratog Carcinog Mutagen, 20:35, 2000 : PubMed
The developmental toxicity of thalidomide was evaluated using FETAX (Frog Embryo Teratogenesis Assay - Xenopus). Young X. Laevis embryos were exposed to this compound in each of two concentration-response experiments with and without differently induced exogenous metabolic activation systems (MASs) and/or inhibited MASs. Young male Sprague-Dawley rats were treated with either isoniazid or Aroclor 1254 to induce cytochrome P-450. Several of the rats were subsequently treated with diethyl maleate (DM) to deplete glutathione reserves. Specific aliquots of rat liver microsomes were treated with 3-amino-1,2,4-triazole (ATZ) or alpha-napthoflavone (alpha-N) to selectively inhibit P-450 activity. Bioactivation was indicated by increased developmental toxicity observed in MAS tests. Results obtained indicated that thalidomide was predominantly activated by P-450 isozyne CYP2E1, although weak cross-specificity between CYP1A1/A2 may have existed. Detoxification pathways for thalidomide were investigated by treatment of the MAS with cyclohexene oxide (CHO) and DM to inhibit the epoxide hydrolase and glutathione conjugation pathways, respectively. Results indicated that epoxide hydrolase was primarily responsible for the detoxification of bioactivated thalidomide. Teratogenesis Carcinog. Mutagen. 20:35-47, 2000.
        
Title: Synergism of tebufenozide in resistant and susceptible strains of obliquebanded leafroller (Lepidoptera: Tortricidae) and resistance to new insecticides Waldstein DE, Reissig WH Ref: J Econ Entomol, 93:1768, 2000 : PubMed
Cross-resistance of the obliquebanded leafroller, Choristoneura rosaceana (Harris), to tebufenozide was reported from laboratory studies before it had been used in commercial orchards in New York State. Bioassays with obliquebanded leafroller larvae from tebufenozide and organophosphate susceptible and resistant colonies were conducted with chlorfenapyr, emamectin benzoate, fenoxycarb, fipronil, spinosad, and tebufenozide to determine if cross-resistance was present before these new insecticides were introduced into commercial orchards. Resistance ratios ranged from 1.1 to 3.2 for all insecticides except tebufenozide (12.8). Significant differences between susceptible and resistant colonies were found with emamectin benzoate, fenoxycarb, and fipronil. The effect of the metabolic synergists piperonyl butoxide (PBO) and diethyl maleate (DEM) on tebufenozide toxicity was examined to determine mechanisms for obliquebanded leafroller resistance to tebufenozide and potential mechanisms for other new insecticides. At a concentration of 20 ppm, PBO and DEM significantly synergized the toxicity of tebufenozide in resistant and susceptible colonies (three- to fourfold). Obliquebanded leafrollers may be resistant to new insecticides with distinct modes of action even if these compounds have not been previously used in commercial orchards.
        
Title: Characterization of malathion resistance in a Mexican population of Rhizopertha dominica Guedes RNC, Zhu KY Ref: Pest Sci, 53:15, 1998 : PubMed
Malathion resistance of a field-collected population of Rhizopertha dominica (Coleoptera: Bostrichidae) from Mexico was evaluated and the resistance mechanisms were characterized both in vivo and in vitro. The Mexican population showed a resistance level of 50-fold at LC50 as compared with that of a susceptible laboratory population. Malathion bioassays with the synergists triphenyl phosphate, piperonyl butoxide and diethyl maleate suggested that esterases were likely to contribute to the resistance whereas cytochrome P450 monooxygenases and glutathione S-transferases were not. In-vitro assays of esterases indicated that the general esterase activity was 1.3-fold higher in the Mexican population than in the susceptible population. However, the phosphotriesterase activity in the resistant population was 3.7-fold higher than in the susceptible population. Significantly higher phosphotriesterase activity in the resistant population was further indicated by 3.4-fold increase of Vmax in enzyme kinetics and higher frequency of individuals with high phosphotriesterase activity in this population. All these findings suggested that phosphotriesterases play a role in malathion resistance in the Mexican population of lesser grain borer.
        
Title: Toxicity of spinosad to susceptible and resistant strains of house flies, Musca domestica Scott JG Ref: Pest Sci, 54:131, 1998 : PubMed
The toxicity of spinosad, a new insecticide derived from the bacterium Saccharopolyspora spinosa, was evaluated against susceptible and resistant strains of house fly (Musca domestica L.). Spinosad was highly toxic to house flies based on 72-h LD50 values and the symptoms of poisoning were consistent with a neurotoxic mechanism of action. Spinosad was relatively slow acting, with the maximum toxicity noted at 72 h. Piperonyl butoxide and S,S,S,-tribu-tylphosphorotrithioate synergized the toxicity of spinosad by 3.0- and 1.8-fold, respectively, while diethyl maleate had no significant effect. These results suggest that there is a small degree of monooxygenase-mediated spinosad detoxification in house flies, while hydrolases may be only minimally important and glutathione transferases may have no role. There were no substantial levels of cross-resistance detected, except in the LPR strain where a low 4.3-fold cross-resistance was observed. The cyclodiene-resistant OCR strain was 2.7-fold more sensitive to spinosad than the susceptible strain (CS). These results suggest that cross-resistance may not be a limiting factor for the use of spinosad against house flies.
        
Title: Effect of insecticide synergist combinations on the survival of Spodoptera exigua Van Laecke K, Degheele D Ref: Pest Sci, 37:283, 1993 : PubMed
The chitin synthesis inhibitors diflubenzuron and teflubenzuron have recently become ineffective for the control of Spodoptera exigua in floricultural crops. An extended laboratory test with second-instar larvae of S. exigua on Vicia faba plants was carried out to determine the influence of synergists on the biological activity of three benzoylphenyl ureas (BPUs). The co-application of piperonyl butoxide, an oxidase inhibitor, did not increase the activity of diflubenzuron, teflubenzuron or hexaflumuron. The best results were obtained with diethyl maleate, for suppressing glutathione S-transferase activity, and with dimethoate, as a hydrolase inhibitor. A joint application of diflubenzuron (at a concentration which resulted in 43% survival) with diethyl maleate or dimethoate gave only 6.2 and 8.9% surviving larvae, respectively. In addition, development to fourth-instar larvae was inhibited. The more stable teflubenzuron was synergized by both compounds to a much lesser extent than diflubenzuron. None of the synergists had a significant effect on the activity of hexaflumuron, which was the most potent insecticide of the three BPUs tested against S. exigua.
        
Title: Differential toxicity of cocaine and its isomers, (+)-cocaine and (-)-psi-cocaine, is associated with stereoselective hydrolysis by hepatic carboxylesterases in cultured rat hepatocytes Melchert RB, Goldlin C, Zweifel U, Welder AA, Boelsterli UA Ref: Chemico-Biological Interactions, 84:243, 1992 : PubMed
Cocaine induces acute lethal cell injury in rat hepatocytes following N-oxidative metabolic activation by cytochrome P450-dependent and flavin-dependent monooxygenases. Beside this oxidative bioactivation pathway, hepatic carboxylesterases may cleave the carboxymethylester or the benzoylester linkage which leads to molecules found to be non-toxic in vivo. To elucidate the structural requirements of the cocaine molecule for its bioactivation and inactivation, the cytotoxic potential of the natural (-)-cocaine relative to two isomeric forms, (+)-cocaine* (the unnatural enantiomer) and (-)-psi-cocaine (the C2 epimer of the unnatural cocaine) were investigated. Primary short-term cultures of rat hepatocytes obtained from phenobarbital (PB)-pretreated rats were exposed to the drugs for up to 24 h. (-)-Cocaine produced marked time- and concentration-dependent release of lactate dehydrogenase (LDH) into the extracellular medium, whereas the other forms were not cytotoxic (0-1 mM). Furthermore, depletion of cellular glutathione (GSH) with diethylmaleate enhanced LDH release in (-)-cocaine-treated cells and caused marginal cytotoxicity in hepatocytes exposed to the other isomers. To investigate the mechanisms that could be responsible for these isomer-specific effects, the time-dependent metabolic degradation was determined both in cultured hepatocytes and in hepatic microsomes in the presence or absence of the serine carboxylesterase inhibitors, phenylmethylsulfonylfluoride (PMSF) or NaF. All three cocaine analogs were enzymatically degraded, but the rates of ester cleavage greatly varied among the stereoisomers. (-)-Cocaine was primarily N-oxidized via SKF-525A-sensitive pathways, whereas (+)-cocaine was predominantly hydrolyzed by PMSF-sensitive carboxylesterases. In contrast, (-)-psi-cocaine, which is very stable in the absence of cells at 37 degrees C and pH 7.4, was subject to extremely fast enzymatic ester cleavage. In conclusion, these results indicate that the isomer-specific differential cytotoxicity of (-)-cocaine, (+)-cocaine and (-)-psi-cocaine in hepatocytes may be related to stereoselective differences in the rates of hydrolytic inactivation by hepatic carboxylesterases and that the N-oxidative pathway, resulting in hepatocyte injury, may thus be relevant only for (-)-cocaine.
        
Title: Synergism of organophosphorus insecticides by diethyl maleate and related compounds in house flies Welling W, de Vries JW Ref: Pesticide Biochemistry and Physiology, 23:358, 1985 : PubMed
The toxicity of several organophosphorus and one carbamate insecticide for house flies is enhanced by simultaneous administration of diethyl maleate. Synergism factors vary from 2 to 116 and are strongly dependent on the combination of strain and insecticide studied. In general, it seems that thiono compounds are less synergized than their oxon analogs. Comparison of some analogs of maleic acid esters showed that the diethyl ester was not the most active compound. trans-Phenylbutenone was found to be a good alternative for diethyl maleate. Both diethyl maleate and trans-phenylbutenone deplete glutathione in the flies, but this depletion per se is certainly not the most important mode of action. In vitro experiments made clear that sufficient glutathione remained to permit a substantial insecticide degradation rate. Moreover, treatment of the flies with diethyl maleate 24 hr before the insecticide application resulted in a partial or complete disappearance of synergism in a period in which the glutathione concentration in the flies was still very low. Apart from this effect on the glutathione levels in the insect tissues it appeared from further experimental work in vitro that both compounds had a direct inhibitory effect on glutathione-dependent transferase(s) as well as on oxidative enzymes involved in insecticide degradation. The contribution of these reactions to the eventual synergism factor is discussed.
        
Title: Parathion and methyl parathion toxicity and metabolism in piperonyl butoxide and diethyl maleate pretreated mice Mirer FE, Levine BS, Murphy SD Ref: Chemico-Biological Interactions, 17:99, 1977 : PubMed