Voltage‐sensitive sodium channel mutations S989P + V1016G in Aedes aegypti confer variable resistance to pyrethroids,DDT and oxadiazines

BACKGROUND

Aedes aegypti is a vector of several important human pathogens. Control efforts rely primarily on pyrethroid insecticides for adult mosquito control, especially during disease outbreaks. A. aegypti has developed resistance nearly everywhere it occurs and insecticides are used. An important mechanism of resistance is due to mutations in the voltage‐sensitive sodium channel (Vssc) gene. Two mutations, in particular, S989P + V1016G, commonly occur together in parts of Asia.

RESULTS

We have created a strain (KDR:ROCK) that contains the Vssc mutations S989P + V1016G as the only mechanism of pyrethroid resistance within the genetic background of Rockefeller (ROCK), a susceptible lab strain. We created KDR:ROCK by crossing the pyrethroid‐resistant strain Singapore with ROCK followed by four backcrosses with ROCK and Vssc S989P + V1016G genotype selections. We determined the levels of resistance conferred to 17 structurally diverse pyrethroids, the organochloride DDT, and oxadiazines (VSSC blockers) indoxacarb (proinsecticide) and DCJW (the active metabolite of indoxacarb). Levels of resistance to the pyrethroids were variable, ranging from 21‐ to 107‐fold, but no clear pattern between resistance and chemical structure was observed. Resistance is inherited as an incompletely recessive trait. KDR:ROCK had a > 2000‐fold resistance to DDT, 37.5‐fold cross‐resistance to indoxacarb and 13.4‐fold cross‐resistance to DCJW.

CONCLUSION

Etofenprox (and DDT) should be avoided in areas where Vssc mutations S989P + V1016G exist at high frequencies. We found that pyrethroid structure cannot be used to predict the level of resistance conferred by kdr. These results provide useful information for resistance management and for better understanding pyrethroid interactions with VSSC. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Frequency and intensity of pyrethroid resistance through the CDC bottle bioassay and their association with the frequency of kdr mutations in Aedes aegypti (Diptera: Culicidae) from Mexico

BACKGROUND

The control of Aedes aegypti (L.), the main urban vector that causes arboviral diseases such as dengue, Chikungunya and Zika, has proved to be a challenge because of a rapid increase in insecticide resistance. Therefore, adequate monitoring of insecticide resistance is an essential element in the control of Ae. aegypti and the diseases it transmits. We estimated the frequency and intensity (Resistance Frequency Rapid Diagnostic Test [F‐RDT] and Resistance Intensity Rapid Diagnostic Test [I‐RDT]) of pyrethroid resistance in populations of Ae. aegypti from Mexico using the bottle bioassay and results were related to the frequencies of knockdown resistance (kdr) mutations V1016I and F1534C.

RESULTS

All populations under study were resistant to the pyrethroids: bifenthrin (99%), d‐(cis–trans)‐phenothrin (6.3% cis, 91.7% trans) and permethrin (99.5%) according to F‐RDT, and showed moderate to high‐intensity resistance at 10× the diagnostic dose (DD) in I‐RDT. Frequencies of the kdr mutation V1016I in Ae. aegypti populations were correlated with moderate permethrin resistance at 10× DD, whereas F1534C mutation frequencies were correlated with high bifenthrin resistance at 5× DD. Both I1016 and C1535 were highly correlated with high‐intensity phenothrin resistance at 1× to 10× DD.

CONCLUSIONS

This study showed that high frequencies of kdr mutations V1016I and F1534C are reflected in the results of F‐RDT and I‐RDT tests. Bioassays in conjunction with the characterization of genetic resistance mechanisms are indispensable in the strategic and rational management of resistance in mosquitoes. © 2018 Society of Chemical Industry

     

Resistance management strategies in malaria vector mosquito control. A large-scale field trial in Southern Mexico

A resistance management programme comparing rotations, mosaics and single use of insecticides for residual house-spraying against the insect vectors of malaria is being carried out in Southern Mexico. The area was chosen because of its prior history of insecticide use, relatively sedentary vector, and physical features of the area which limit inward migration of insects to the study area. A high level of resistance to DDT and low levels of organophosphorus (OP), carbamate and pyrethroid resistance were detected by WHO discriminating-dose assays in field populations of Anopheles albimanus in the pre-spray period in the region where this resistance management project is being undertaken. After the first year of spraying, resistance, as measured by a discriminating-dose assay, was still at a high level for DDT and had risen for all the other insecticides. Biochemical assays showed that DDT resistance was primarily caused by elevated levels of glutathione S-transferase (GST) activity leading to increased rates of metabolism of DDT to DDE. The numbers of individuals with elevated GST and DDT resistance were well correlated, suggesting that this is the only major DDT resistance mechanism in this population. The carbamate resistance in this population was conferred by an altered acetylcholinesterase (AChE) mechanism. The level of resistance in bioassays correlated well with the frequency of individuals homozygous for the altered AChE allele. This suggests that the level of resistance conferred by this mechanism in its heterozygous state is below the level of detection of the bioassay. The low levels of OP and pyrethroid resistance could be conferred by either the elevated esterase or monooxygenase enzymes. The esterases, however, are elevated only with p-nitrophenyl acetate (PNPA), and are unlikely to be causing broad-spectrum OP resistance. The altered AChE mechanism may also be contributing to the OP but not the pyrethroid resistance. There were significant differences in some resistance gene frequencies for insects obtained by different indoor and outdoor trapping methods. To determine whether the different sampling methods were effectively sampling the same interbreeding population, RAPD analysis of insects obtained by different collection methods in different villages was undertaken. There was no observed variability in the RAPD patterns for the different mosquito samples with a number of primers. ©1997 SCI     

Homology modelling of Drosophila cytochrome P450 enzymes associated with insecticide resistance

BACKGROUND: Overexpression of the cytochrome P450 gene Cyp6g1 confers resistance against DDT and a broad range of other insecticides in Drosophila melanogaster Meig. In the absence of crystal structures of CYP6G1 or complexes with its substrates, structural studies rely on homology modelling and ligand docking to understand P450–substrate interactions. RESULTS: Homology models are presented for CYP6G1, a P450 associated with resistance to DDT and neonicotinoids, and two other enzymes associated with insecticide resistance in D. melanogaster, CYP12D1 and CYP6A2. The models are based on a template of the X‐ray structure of the phylogenetically related human CYP3A4, which is known for its broad substrate specificity. The model of CYP6G1 has a much smaller active site cavity than the template. The cavity is also ‘V’‐shaped and is lined with hydrophobic residues, showing high shape and chemical complementarity with the molecular characteristics of DDT. Comparison of the DDT–CYP6G1 complex and a non‐resistant CYP6A2 homology model implies that tight‐fit recognition of this insecticide is important in CYP6G1. The active site can accommodate differently shaped substrates ranging from imidacloprid to malathion but not the pyrethroids permethrin and cyfluthrin. CONCLUSION: The CYP6G1, CYP12D1 and CYP6A2 homology models can provide a structural insight into insecticide resistance in flies overexpressing P450 enzymes with broad substrate specificities. Copyright © 2010 Society of Chemical Industry     

Mutations in the sodium channel associated with pyrethroid resistance in the greenhouse whitefly, Trialeurodes vaporariorum

BACKGROUND: Trialeurodes vaporariorum Westwood is an important pest of protected crops in temperate regions of the world. Resistance to pyrethroid insecticides is long established in this species, but the molecular basis of the mechanism(s) responsible has not previously been disclosed. RESULTS: Mortality rates of three European strains of T. vaporariorum to the pyrethroid bifenthrin were calculated, and each possessed significant resistance (up to 662‐fold) when compared with a susceptible reference strain. Direct sequencing revealed three amino acid substitutions in the para‐type voltage‐gated sodium channel (the pyrethroid and DDT target site) of bifenthrin‐resistant T. vaporariorum at positions previously implicated with pyrethroid or DDT resistance (M918L, L925I and T929I) in other related species. CONCLUSION: This study indicates that resistance to bifenthrin in T. vaporariorum is associated with target‐site insensitivity, and that the specific mutations in the sodium channel causing resistance may differ between localities. Copyright © 2012 Society of Chemical Industry     

Resistance of Pakistani field populations of spotted bollworm Earias vittella (Lepidoptera: Noctuidae) to pyrethroid,organophosphorus and new chemical insecticides

BACKGROUND: The spotted bollworm Earias vittella (Fab.) is a serious pest of cotton and okra in Pakistan. Owing to persistent use of insecticides, this pest has developed resistance, especially to pyrethroids. The present studies aimed at determining the extent of resistance to pyrethroid, organophosphorus and new chemical insecticides in Pakistani populations of E. vittella. RESULTS: Field populations of E. vittella were monitored at Multan, Pakistan, from 1999 to 2007 for their resistance against six pyrethroid, four organophosphorus and six new chemical insecticides using a leaf‐dip bioassay. Of the pyrethroids, resistance was generally low to zeta‐cypermethrin and moderate to high or very high to cypermethrin, deltamethrin, esfenvalerate, bifenthrin and lambda‐cyhalothrin. Resistance to organophosphates chlorpyrifos, profenofos, triazophos and phoxim was recorded at very low to low levels. Among new chemicals, E. vittella had no or a very low resistance to spinosad, emamectin benzoate and methoxyfenozide, a very low to low resistance to abamectin, a very low to moderate resistance to indoxacarb and a moderate resistance to chlorfenapyr. CONCLUSION: The results indicate a lack of cross‐resistance between pyrethroid and organophosphorus insecticides in E. vittella. Rotation of insecticides showing no, very low or low resistance, but belonging to different insecticide classes with unrelated modes of action, may prevent or mitigate insecticide resistance in E. vittella. Copyright © 2009 Society of Chemical Industry     

Molecular analysis of pyrethroid resistance conferred by target insensitivity and increased metabolic detoxification in Plutella xylostella

BACKGROUND: The pyrethroid resistance of the diamondback moth Plutella xylostella (L.) is conferred by increased gene expression of cytochrome P450 to detoxify the insecticide and/or through gene mutation of the sodium channel, which makes the individual insensitive to pyrethroids. However, no information is available about the correlation between the increased metabolic detoxification and the target insensitivity in pyrethroid resistance. RESULTS: Frequencies of pyrethroid‐resistant alleles (L1014F, T929I and M918I) and two resistance‐related mutations (A1101T and P1879S) at the sodium channel and expression levels of the cytochrome P450 gene CYP6BG1 were examined individually using laboratory and field strains of P. xylostella. Real‐time quantitative PCR analysis using the laboratory strains revealed that levels of larval expression of the resistant strain, homozygous for the pyrethroid‐resistant alleles other than the M918I, are significantly higher than those of the susceptible strain. In the field strains, the expression levels in insects having the same resistant alleles as those of the resistant strains varied greatly among individuals. The expression levels were not significantly higher than those in the heterozygotes. CONCLUSION: Significant correlation between the target insensitivity and the increased metabolic detoxification in pyrethroid resistance of P. xylostella was observed in the laboratory but not in the field. Copyright © 2010 Society of Chemical Industry     

Insecticide resistance compromises the control of Aedes aegypti in Bangladesh

BACKGROUND

With no effective drugs or widely available vaccines, dengue control in Bangladesh is dependent on targeting the primary vector Aedes aegypti with insecticides and larval source management. Despite these interventions, the dengue burden is increasing in Bangladesh, and the country experienced its worst outbreak in 2019 with 101 354 hospitalized cases. This may be partially facilitated by the presence of intense insecticide resistance in vector populations. Here, we describe the intensity and mechanisms of resistance to insecticides commonly deployed against Ae. aegypti in Dhaka, Bangladesh.

RESULTS

Dhaka Ae. aegypti colonies exhibited high-intensity resistance to pyrethroids. Using CDC bottle assays, we recorded 2–24% mortality (recorded at 24 h) to permethrin and 48–94% mortality to deltamethrin, at 10× the diagnostic dose. Bioassays conducted using insecticide–synergist combinations suggested that metabolic mechanisms were contributing to pyrethroid resistance, specifically multi-function oxidases, esterases, and glutathione S-transferases. In addition, kdr alleles were detected, with a high frequency (78–98%) of homozygotes for the V1016G mutation. A large proportion (≤ 74%) of free-flying and resting mosquitoes from Dhaka colonies survived exposure to standard applications of pyrethroid aerosols in an experimental free-flight room. Although that exposure affected the immediate host-seeking behavior of Ae. aegypti, the effect was transient in surviving mosquitoes.

CONCLUSION

The intense resistance characterized in this study is likely compromising the operational effectiveness of pyrethroids against Ae. aegypti in Dhaka. Switching to alternative chemical classes may offer a medium-term solution, but ultimately a more sustainable and effective approach to controlling dengue vectors is required. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Biotype and insecticide resistance status of the whitefly Bemisia tabaci from China

BACKGROUND: Resistance to numerous insecticide classes in Bemisia tabaci Gennadius has impaired field control efficacy in south‐eastern China. The biotype and resistance status of B. tabaci collected from these areas was investigated. RESULTS: Two different biotypes of B. tabaci (B‐biotype and Q‐biotype) were detected in south‐eastern China, and the samples collected from geographical regions showed a prevalence of the Q‐biotype and the coexistence of B‐ and Q‐biotypes in some regions. Moderate to high levels of resistance to two neonicotinoids were established in both biotypes (28–1900‐fold to imidacloprid, 29–1200‐fold to thiamethoxam). Medium to high levels of resistance to alpha‐cypermethrin (22–610‐fold) were also detected in both biotypes. Four out of 12 populations had low to medium levels of resistance to fipronil (10–25‐fold). Four out of 12 populations showed low levels of resistance to spinosad (5.7–6.4‐fold). All populations tested were susceptible to abamectin. CONCLUSION: The Q‐biotype B. tabaci is supplanting the B‐biotype which used to be ubiquitous in China. Field populations of both B‐ and Q‐biotypes of B. tabaci have developed high levels of resistance to imidacloprid and thiamethoxam. Abamectin is the most effective insecticide against adult B. tabaci from all populations. Copyright © 2010 Society of Chemical Industry     

Mechanisms of DDT resistance in larvae of the mosquito Aedes aegypti L

Larvae of eight strains of Aedes aegypti were exposed to DDT and compared for resistance, DDT uptake, in-vivo breakdown of DDT and residual unmetabolised DDT. Resistance varied widely between strains, three being fully susceptible, two almost immune and three of intermediate resistance. Breakdown of DDT by dehydrochlorination to 1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene (pp'-DDE) occurred in all strains and was greater in the five resistant types, but there was no significant correlation between the extent of breakdown in the resistant strains and the level of resistance. Moreover the overall difference between susceptible and resistant strains disappeared when they were compared at a low, almost sublethal, concentration of DDT. Larvae of resistant strains carried a greater absolute quantity of unmetabolised DDT in the body and were able to tolerate levels of DDT that were lethal to susceptible larvae. However the two most resistant strains (T8 and B51) contained significantly less DDT plus pp'-DDE than strains of intermediate resistance (T30 and BSJ) from which they had been derived. Addition of the synergist chlorfenethol to DDT increased its knockdown effect on all resistant strains, suggesting that dehydrochlorination was a factor in resistance. Three strains, two DDT-resistant and one DDT-susceptible, were tested with 1,1-bis(4-ethoxyphenyl)-2,2-dimethylpropane (I), an insecticide that cannot be dehydrochlorinated. All the strains were relatively tolerant to it although the DDT-susceptible strains were less tolerant. Addition of the synergist sesamex decreased the level of tolerance to I in all strains which suggested that microsomal oxidation made some contribution to it. It is concluded that three factors contribute to larval DDT resistance in A. aegypti; (a) increased metabolism to pp'-DDE; (b) increased tolerance to unmetabolised internal DDT; and (c) reduced content of DDT+pp'-DDE (only in the most resistant strains and due either to reduced absorption or increased excretion). These factors are discussed in relation to known larval resistance genes R^DDT1 and y.     

The pyrethroid resistance status and mechanisms in Aedes aegypti from the Guerrero state,Mexico

Dengue is one of the most important vector-borne diseases worldwide and is a public health problem in Mexico. Most programs in dengue endemic countries rely on insecticides for Aedes control. In Mexico, pyrethroid insecticides (mainly permethrin and deltamethrin) have been extensively used over a decade as adulticides and represented a strong selection for insecticide resistance for dengue vectors in several parts of the country. We studied the type, frequency and distribution of insecticide resistance mechanisms in Aedes aegypti from six municipalities in the state of Guerrero selected on the basis of historically intense chemical control and a high risk for dengue transmission. Ae. aegypti eggs were collected from October 2009 to January 2010 using ovitraps. F₁ adults, emerged from these collections, were exposed to permethrin, deltamethrin and DDT in WHO diagnostic tests and showed high resistance levels to both pyrethroids and DDT. This was consistent with the presence of increased metabolic enzyme activities and target site insensitivity due to kdr mutations. Biochemical assays showed elevated esterase and glutathione S-transferase activities in the six municipalities. The V1016I kdr mutation on the IIS6 domain of the sodium channel gene was present in an overall frequency of 0.80. A second mutation, F1534C on the IIIS6 domain of the same gene was also detected, being the first report of this mutation in Guerrero. The multiple resistance mechanisms present in Ae. aegypti from Guerrero state represent a warning for the efficacy of the pyrethroid usage and consequently for the success of the dengue control program.     

Occurrence of insecticide resistant pollen beetles (Meligethes aeneus F.) in Danish oilseed rape (Brassica napus L.) crops

The pollen beetle is the most important pest in Danish oilseed rape crops and it is essential that farmers are able to control this pest, especially in years when the economic damage threshold will be exceeded. About seven years ago, Danish oilseed rape growers began reporting that the pyrethroids seemed to be losing effectiveness towards pollen beetles. In 2001 18 populations collected from winter and spring oilseed rape fields were tested and very high level of insecticide resistance was found. In 2003 100 populations of pollen beetles were tested, using a FAO dip-test, for resistance to fenitrothion, lambda-cyhalothrin and tau-fluvalinate. It can be concluded that many Danish populations of pollen beetles are resistant to the pyrethroid lambda-cyhalothrin and to a lesser extent to the pyrethroid tau-fluvalinate. No resistance to the organophosphate fenitrothion was found.     

Pyrethroids, knockdown resistance and sodium channels

Knockdown resistance to DDT and the pyrethrins was first described in 1951 in the housefly (Musca domestica L.). This trait, which confers reduced neuronal sensitivity to these insecticides, was subsequently shown to confer cross-resistance to all synthetic pyrethroid insecticides that have been examined to date. As a consequence, the worldwide commercial development of pyrethroids as a major insecticide class over the past three decades has required constant awareness that pyrethroid overuse has the potential to reselect this powerful resistance mechanism in populations that previously were resistant to DDT. Demonstration of tight genetic linkage between knockdown resistance and the housefly gene encoding voltage-sensitive sodium channels spurred efforts to identify gene mutations associated with knockdown resistance and understand how these mutations confer a reduction in the sensitivity of the pyrethroid target site. This paper summarizes progress in understanding pyrethroid resistance at the molecular level, with particular emphasis on studies in the housefly.     

Imidacloprid‐susceptible Nilaparvata lugens individuals exceeded resistant individuals in a mixture population with density pressure

BACKGROUND

Fitness costs associated with insecticide resistance in pest insects have mainly been studied under optimal laboratory conditions. However, resistant insects face more stressors than just insecticides in the field, and how the resistant population reacts to these stressors is of practical importance for the control of pest insects such as the brown planthopper Nilaparvata lugens. The aim of the present study was to explore the impact of population density on the competitiveness of resistant and susceptible individuals.

RESULTS

Two isogenic N. lugens populations, a highly imidacloprid‐resistant population (HZ‐R) with a resistance ratio (RR) of 227.10 and a relatively susceptible population (HZ‐S) with an RR of 2.99, were created from a field‐resistant population (HZ; RR 62.51). The high resistance levels of HZ‐R and HZ were mainly attributable to the overexpression of multiple cytochrome P450 (CYP) genes such as CYP6ER1, CYP6AY1, CYP6CW1 and CYP4CE1 compared with HZ‐S, this being supported by piperonyl butoxide synergism. HZ‐R was observed to be more resistant to thiacloprid and etofenprox compared with HZ and HZ‐S. Most interestingly, in high population density treatments, HZ‐S individuals were much more competitive than HZ‐R individuals.

CONCLUSION

Imidacloprid‐resistant individuals of N. lugens are less competitive than their susceptible counterparts under density pressure. © 2017 Society of Chemical Industry     

Over-expression of cytochrome P450 CYP6B7 mRNA and pyrethroid resistance in Australian populations of Helicoverpa armigera (Hübner)

Three cDNA clones for cytochrome P450s, CYP6B2, CYP6B6 and CYP6B7 which have 84–87% protein sequence identity have been isolated previously from Helicoverpa armigera, and the CYP6B7 mRNA was found to be over-expressed in a pyrethroid-resistant field population. Subsequent analysis has shown that over-expression is observed in a majority of individuals in all populations tested. Single-pair crosses between resistant and sensitive individuals indicated that the pyrethroid resistance co-segregated with the over-expression of this mRNA. Southern analysis indicated that the over-expression, which was confined to midgut only in many insects, was not related to gene amplification. These observations add weight to the conclusion that CYP6B7 is the cytochrome P450 form involved in pyrethroid resistance, and that over-expression of cytochrome P450 CYP6B7 is a common cause of pyrethroid resistance in H. armigera. The results suggest that specific probes for CYP6B7 protein or mRNA could provide the basis for the development of tools for monitoring pyrethroid resistance due to mixed function oxidase activity in field populations of this insect. © 1998 Society of Chemical Industry     

Dynamics of pyrethroid resistance in a field population of Helicoverpa armigera (Hübner) in China

Dynamics of pyrethroid resistance in a field population of cotton bollworm (Helicoverpa armigera) was demonstrated by continuous monitoring with twin discriminating dosages, and the influencing factors were also experimentally analysed. Resistance in a field population in China increased rapidly in the 3rd and 4th generations when population density became higher and insecticides were applied repeatedly, then decreased suddenly during over-wintering and slowly in the 1st and 2nd generations when insecticide spraying was suspended. Resistance increase could be countered by dilution as a result of immigration of susceptible moths from corn fields, which were found to be a natural refuge for this pest in China. The reduction of resistance during over-wintering and the 1st and 2nd generations was affected by the lower fitness of resistant cotton bollworms to low temperature and disadvantages in reproduction. The possibilities of managing the resistance in field populations on the basis of these observations are discussed. © 1999 Society of Chemical Industry     

Pyrethroid resistance in the tomato red spider mite, Tetranychus evansi, is associated with mutation of the para-type sodium channel

BACKGROUND: The tomato red spider mite, Tetranychus evansi (Baker and Pritchard), is a serious pest of solanaceous crops in many African countries. In this study an investigation has been conducted to establish whether mutation of the para‐type sodium channel underlies pyrethroid resistance in T. evansi strains collected in Southern Malawi. RESULTS: Two T. evansi strains from Malawi showed tolerance to the organophosphate chlorpyrifos and resistance (20–40‐fold) to the pyrethroid bifenthrin, but were susceptible to two contemporary acaricides (abamectin and fenpyroximate) in insecticide bioassays. Cloning of a 3.1 kb fragment (domains IIS5 to IVS5) of the T. evansi para gene from pyrethroid‐resistant and pyrethroid‐susceptible strains revealed a single non‐synonymous mutation in the resistant strains that results in an amino acid substitution (M918T) within the domain II region of the channel. Although novel to mites, this mutation confers high levels of resistance to pyrethroids in several insect species where it has always been associated with another mutation (L1014F). This is the first report of the M918T mutation in the absence of L1014F in any arthropod species. Diagnostic tools were developed that allow sensitive detection of this mutation in individual mites. CONCLUSION: This is the first study of pyrethroid resistance in T. evansi and provides contemporary information for resistance management of this pest in Southern Malawi. Copyright © 2011 Society of Chemical Industry     

Insecticide resistance profiles can be misleading in predicting the survival of Myzus persicae genotypes on potato crops following the application of different insecticide classes

BACKGROUND: The accuracy of predicting the survival of insecticide‐resistant aphids following the application of commonly used insecticides from the carbamate, the pyrethroid, a mix of the two or the neonicotinoid chemical classes was evaluated in a potato field in Scotland. Equal proportions of five genotypes of the peach‐potato aphid, Myzus persicae (Sulzer), with none, resistance to dimethyl‐carbamates, resistance to pyrethroids or combinations conferring resistance to both chemical classes were released into potato field plots. The insecticides were sprayed separately onto these plots, the aphid populations were analysed after 6–8 days and the process repeated. RESULTS: For each assessment after the three separate spray events, plots treated with the carbamate had 48, 147 and 28%, those treated with pyrethroid 53, 210 and 89%, those treated with carbamate/pyrethroid 28, 108 and 64% and those treated with neonicotinoid 43, 55 and 11% of the numbers of M. persicae by comparison with untreated controls. Only the proportions of surviving aphids from the genotype containing no insecticide resistance traits and the genotype containing elevated carboxylesterases matched ratios predicted from the selective advantage afforded by the resistance traits alone. Survival of aphids from the other three genotypes that carried 1–3 of the insecticide resistance traits differed from expectations in all cases, possibly owing to physiological differences, including their vulnerability to predators and hymenopterous parasitoids present at the site and/or their carrying unknown insecticide resistance mechanisms. CONCLUSION: Control strategies based on knowledge of the genetically determined insecticide resistance profile of an M. persicae population alone are insufficient. Hence, other important factors contributing to aphid survival under insecticide pressure need to be considered. Copyright © 2012 Society of Chemical Industry     

Cloning and characterization of two glutathione S-transferases from pyrethroid-resistant Culex pipiens

BACKGROUND: The Marin strain of Culex pipiens Say is a pyrethroid‐resistant population that was collected in Marin County, California, in 2001 and subsequently maintained in the laboratory under regular permethrin exposure. RESULTS: In this study, two cDNAs, CpGSTd1 and CpGSTd2, encoding glutathione S‐transferase (GST) were cloned from Cx. pipiens Marin. Phylogenetic analysis of the deduced amino acid sequences, CpGSTD1 and CpGSTD2, of these genes indicated that they belong to the Delta class of insect GSTs. The nucleotide and deduced amino acid sequences of CpGSTd1 and CpGSTd2 were 59 and 48% identical respectively. CpGSTD1 and CpGSTD2 were expressed in Escherichia coli and purified by affinity chromatography. The recombinant GSTs exhibited unique selectivity towards the general GST substrates 1‐chloro‐2,4‐dinitrobenzene (CDNB) and 1,2‐dichloro‐4‐nitrobenzene (DCNB), and also differed in their sensitivity to known inhibitors of GSTs. CpGSTD1 exhibited peroxidase activity with cumene hydroperoxide, while CpGSTD2 appeared to lack this activity. CpGSTD1 was able to metabolize 1,1,1‐trichloro‐2,2‐bis(4‐chlorophenyl)ethane (DDT), while DDT metabolism by CpGSTD2 was not detectable. CpGSTD1 and CpGSTD2 showed no detectable metabolism of permethrin. Gene expression of CpGSTd1 and CpGSTd2 in Marin mosquitoes was elevated about twofold in comparison with that found in a pyrethroid‐sensitive mosquito strain. CONCLUSION: The results indicate that CpGSTD1 and CpGSTD2 have unique biochemical characteristics, but they do not appear to play major roles in permethrin resistance in Marin mosquitoes. Copyright © 2012 Society of Chemical Industry     

Larvicidal activity of Cnidium monnieri fruit coumarins and structurally related compounds against insecticide-susceptible and insecticide-resistant Culex pipiens pallens and Aedes aegypti

BACKGROUND: An assessment was made of the toxicity of imperatorin and osthole identified in Cnidium monnieri fruit, 11 related compounds and five insecticides to larvae from insecticide‐susceptible Culex pipiens pallens (KS‐CP strain) and Aedes aegypti and wild C.p. pallens (YS‐CP colony) using a direct‐contact mortality bioassay. Results were compared with those of the conventional larvicide temephos. RESULTS: Imperatorin (LC₅₀ = 3.14 and 2.88 mg L^?1) was 1.9‐, 3.7‐ and 4.2‐fold and 2.4‐, 4.5‐ and 4.6‐fold more toxic than isopimpinellin, isoimperatorin and osthole against susceptible C. p. pallens and A. aegypti larvae respectively. Overall, all of the compounds were less toxic than temephos (0.011 and 0.019 mg L^?1). The toxicity of these compounds was virtually identical against larvae from the two Culex strains, even though YS‐CP larvae were resistant to fenthion (resistance ratio RR = 390), deltamethrin (RR = 164), cyfluthrin (RR = 14) and temephos (RR = 14). This finding indicates that the coumarins and the insecticides do not share a common mode of action. The structure–activity relationship indicates that the chemical structure and alkoxy substitution and length of the alkoxyl side chain at the C8 position are essential for imparting toxicity. CONCLUSION: The C. monnieri fruit‐derived coumarins and the related coumarins described merit further study as potential insecticides or lead molecules for the control of insecticide‐resistant mosquito populations. Copyright © 2012 Society of Chemical Industry