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1.
Head lice resistance to permethrin is mainly conferred by the knockdown resistance (kdr) trait, a voltage-sensitive sodium channel (VSSC) insensitivity factor. Three VSSC mutations (M815I, T917I and L920F) have been identified. Functional analysis of the mutations using the house fly VSSC expressed in Xenopus oocytes revealed that the permethrin sensitivity is reduced by the M827I (M815I) and L932F (L920F) mutations when expressed alone but virtually abolished by the T929I (T917I) mutation, either alone or in combination. Thus, the T917I mutation is primarily responsible for permethrin resistance in head lice. Comparison of the expression rates of channel variants indicates that the M815I mutation may play a role in rescuing the decreased expression of channels containing T917I. A step-wise resistance monitoring system has been established based on molecular resistance detection techniques. Quantitative sequencing (QS) has been developed to predict the VSSC mutation frequency in head lice at a population basis. The speed, simplicity and accuracy of QS made it an ideal candidate for a routine primary resistance monitoring tool to screen a large number of wild louse populations as an alternative to conventional bioassay. As a secondary monitoring method, real-time PASA (rtPASA) has been devised for more precise determination of low resistance allele frequencies. To obtain more detailed information on resistance allele zygosity, as well as allele frequency, serial invasive signal amplification reaction (SISAR) has been developed as an individual genotyping method. Our approach of using three tiers of molecular resistance detection should facilitate large-scale routine resistance monitoring of permethrin resistance in head lice using field-collected samples.  相似文献   

2.
The resistance levels of different human head louse populations from the USA to 1% permethrin were evaluated using permethrin-impregnated, filter paper disk-contact bioassay. Populations from southern California, south Florida and south central Texas showed 1.5-, 3.1-, and 1.5- to 5.1-fold resistance compared to insecticide-susceptible head louse populations from Panama or Ecuador. Permethrin-resistant or permethrin-susceptible homozygous or heterozygous genotypes were determined from sequences of PCR-amplified genomic DNA fragments of the voltage-sensitive sodium channel α-subunit gene by the presence of a T or C, or both, respectively, at nucleotide positions 36 and 44 in the sequence. The presence of a T at both these positions resulted in the amino acid substitutions, T929I and L932F, respectively. Of the 424 louse samples examined that had the T929I mutation, all also possessed the L932F mutation, indicating that the two mutations were tightly linked. The southern California population was phenotypically determined by bioassay to be comprised of 45% resistant individuals and had a resistant allele frequency of 0.53 by DNA sequence analysis. The south Florida population was phenotypically determined to consist of 87% resistant individuals and had a resistant allele frequency of 0.97. The four Texas populations varied in the level of resistance and in resistant allele frequency. The Mathis population was phenotypically determined to consist of 15% resistant individuals and had a resistant allele frequency of 0.33. However, the populations from San Antonio, Mansfield, and Corpus Christi were likewise phenotyped to have 91%, 94%, and 100%, respectively, resistant individuals and a 0.98, 1.00 and 1.00, respectively, resistant allele frequency. The log survival time versus logit mortality regression lines of susceptible-homozygotes, resistant-homozygotes, and heterozygotes determined that the resistance trait was complete recessive. Thus, the presence of homozygotes of the T929I and L932F mutations in the voltage-sensitive sodium channel correlated well with increased survival time following exposure to permethrin and indicates that a knockdown-type nerve insensitivity mechanism is functioning as the major mechanism causing permethrin resistance in USA head louse populations. Our results substantiate that permethrin resistance in human head louse population in the USA is widespread but variable. Permethrin resistance is highly correlated with the presence of the T929I and L932F point mutations, which are suitable for detection by a variety of DNA-based diagnostic techniques [Pest Manag. Sci. 57 (2001) 968]. Large-scale monitoring of permethrin resistance is possible utilizing these techniques and would provide critical information necessary for the development of an effective resistance management program for pediculosis.  相似文献   

3.
Permethrin resistance in the human head louse, Pediculus capitis De Geer (Anopulura: Pediculidae), has been reported worldwide, is associated with the knockdown phenotype, and elicits cross-resistance to DDT and the pyrethrins. Two point mutations, T929I and L932F, in the voltage-sensitive sodium channel α-subunit gene are responsible for permethrin resistance as a resistant haplotype (kdr-like). We have optimized a serial invasive signal amplification reaction (SISAR) protocol for the detection of these mutations using PCR amplified DNA fragments. SISAR distinguished all genotypes with high accuracy in a head louse population from Texas that was heterogeneous in terms of permethrin sensitivity. Using SISAR, resistance-conferring mutations are detected in a high throughput format, facilitating the efficient monitoring of permethrin resistance allele frequency in field populations.  相似文献   

4.
BACKGROUND: Pediculosis is the most prevalent parasitic infestation of humans. Resistance to pyrethrin‐ and pyrethroid‐based pediculicides is due to knockdown (kdr)‐type point mutations in the voltage‐sensitive sodium channel α‐subunit gene. Early detection of resistance is crucial for the selection of effective management strategies. RESULTS: Kdr allele frequencies of lice from 14 countries were determined using the serial invasive signal amplification reaction. Lice collected from Uruguay, the United Kingdom and Australia had kdr allele frequencies of 100%, while lice from Ecuador, Papua New Guinea, South Korea and Thailand had kdr allele frequencies of 0%. The remaining seven countries investigated, including seven US populations, two Argentinian populations and populations from Brazil, Denmark, Czech Republic, Egypt and Israel, displayed variable kdr allele frequencies, ranging from 11 to 97%. CONCLUSION: The newly developed and validated SISAR method is suitable for accurate monitoring of kdr allele frequencies in head lice. Proactive management is needed where kdr‐type resistance is not yet saturated. Based on sodium channel insensitivity and its occurrence in louse populations resistant to pyrethrin‐ and pyrethroid‐based pediculicides, the T917I mutation appears to be a key marker for resistance. Results from the Egyptian population, however, indicate that phenotypic resistance of lice with single or double mutations (M815I and/or L920F) should also be determined. Copyright © 2010 Society of Chemical Industry  相似文献   

5.
Tetranychus urticae Koch is the most serious mite pest to various orchard trees and garden plants. Biochemical and molecular analyses were conducted to elucidate resistance mechanisms in a fenpropathrin-resistant mite strain (FenR). No significant differences were found in the activities of carboxylesterase and glutathione-S-transferase between the susceptible (UD and PyriF) and FenR strains. Cytochrome P450 activity was highest in PyriF, followed by FenR and UD. Analysis of detoxification enzyme assays, therefore, suggested that metabolic detoxification plays little role, if any, in fenpropathrin resistance. However, the FenR strain showed approximately 104- and 33.3-fold slower knockdown responses than UD and PyriF strains, respectively, suggestive of sodium channel insensitivity as a major resistance mechanism. We cloned cDNA fragments of the para-homologous sodium channel α-subunit gene (Tuvssc) and determined its full-length nucleotide sequences. The complete open reading frame of Tuvssc was 6627 nucleotides, encoding 2209 amino acids. The amino acid sequences of Tuvssc were 47.5% and 51.2% identical to the fruit fly and varroa mite, respectively. Amino acid sequence comparison between the three strains revealed two mutations (L1022V and A1376D) and one deletion (HisDel1278-1280) found only in FenR mites, among which the L1022V mutation was proposed to play a major role in knockdown resistance to fenpropathrin.  相似文献   

6.
Resistance in a dual malathion- and permethrin-resistant head louse strain (BR-HL) was studied. BR-HL was 3.6- and 3.7-fold more resistant to malathion and permethrin, respectively, compared to insecticide-susceptible EC-HL. S,S,S-Tributylphosphorotrithioate synergized malathion toxicity by 2.1-fold but not permethrin toxicity in BR-HL. Piperonyl butoxide did not synergize malathion or permethrin toxicity. Malathion carboxylesterase (MCE) activity was 13.3-fold and general esterase activity was 3.9-fold higher in BR-HL versus EC-HL. There were no significant differences in phosphotriesterase, glutathione S-transferase, and acetylcholinesterase activities between strains. There was no differential sensitivity in acetylcholinesterase inhibition by malaoxon. Esterases from BR-HL had higher affinities and hydrolysis efficiencies versus EC-HL using various naphthyl-substituted esters. Protein content of BR-HL females and males was 1.6- and 1.3-fold higher, respectively, versus EC-HL adults. Electrophoresis revealed two esterases with increased intensity and a unique esterase associated with BR-HL. Thus, increased MCE activity and over-expressed esterases appear to be involved in malathion resistance in the head louse.  相似文献   

7.
To investigate the level of pyrethroid resistance in Anopheles sinensis Wiedemann 1828 (Diptera: Culicidae), a major malaria vector in Korea, we cloned and sequenced the IIS4-6 transmembrane segments of the sodium channel gene that encompass the most widely known kdr mutation sites. Sequence analysis revealed the presence of the major Leu-Phe mutation and a minor Leu-Cys mutation at the same position in permethrin-resistant field populations of An. sinensis. To establish a routine method for monitoring resistance, we developed a simple and accurate real-time PCR amplification of specific allele (rtPASA) protocol for the estimation of resistance allele frequencies on a population basis. The kdr allele frequency of a field population predicted by the rtPASA method (60.8%) agreed well with that determined by individual genotyping (61.7%), demonstrating the reliability and accuracy of rtPASA in predicting resistance allele frequency. Using the rtPASA method, the kdr allele frequencies in several field populations of An. sinensis were determined to range from 25.0 to 96.6%, suggestive of widespread pyrethroid resistance in Korea.  相似文献   

8.
Feng YN  Zhao S  Sun W  Li M  Lu WC  He L 《Pest management science》2011,67(8):904-912
BACKGROUND: The carmine spider mite (CSM), Tetranychus cinnabarinus, is the most harmful mite pest of various crops and vegetable plants. Pyrethroid insecticide fenpropathrin has been used to control insects and mites worldwide, but CSM has developed resistance to this compound. RESULTS: Three synergists together eliminated about 50% resistance against fenpropathrin in the CSM. A point mutation was identified from the sodium channel gene of fenpropathrin‐resistant CSM (FeR) by comparing cDNA sequences between FeR and susceptible (S) sodium channel genes, which caused a phenylalanine (F) to isoleucine (I) change at amino acid 1538 position in IIIS6 of the sodium channel and has been proven to confer strong resistance to pyrethroid in other species. The mRNA expression of the sodium channel gene in the FeR and abamectin‐resistant strain (AbR), which was included as a control, were both relatively lower than in the S. CONCLUSION: These results demonstrate that a mutation (F1538I) is present in the sodium channel gene in FeR of CSM, likely playing an important role in fenpropathrin resistance in T. cinnabarinus, but that decrease in the abundance of sodium channel did not confer this resistance. The F1538I mutation could be used as a molecular marker for detecting kdr resistance in Arachnida populations. Copyright © 2011 Society of Chemical Industry  相似文献   

9.
For testing the susceptibility of the head louse to insecticides impregnated bunches of polyamide fibres, whose diameter resembled that of human hair (0.1 mm) were used. In this manner a low mortality of control lice collected from infected persons was obtained (2.9% after 16 hours of exposure). Even after 25 years of DDT use no apparent resistance could be demonstrated in 25 tested louse populations and cross resistance to permethrin in 7 tested populations. The diagnostic concentration for resistance to pp'-DDT was established at 1%. Of the tested insecticides the highest toxicity was shown by malathion (LC50 = 0.000 018%), pirimiphos-methyl had on average a twice lower toxicity, trichlorphon 104 times lower, tetramethrin 1220 times lower, permethrin 2.5 times lower and pp'-DDT 630 times lower toxicity.  相似文献   

10.
A point mutation in thepara-homologous sodium channel gene has been shown to be associated with knockdown resistance (kdr) in several insect species including the German cockroach. In this study, we analyzed the genomic organization of the region where thekdrmutation resides and then performed polymerase chain reaction (PCR) and sequencing using genomic DNA as the template to detectkdrmutation in 24 pyrethroid-resistant German cockroach strains, most of which have been collected recently from the field. Thekdrmutation, G to C at nt 2979 resulting in a leucine to phenylalanine amino acid substitution, was detected in 20 strains including 2 strains from overseas (China and Germany). Our results clearly indicate that thekdrmutation is widespread in German cockroach populations. However, the super-kdrmutation detected in super-kdrhouse flies was not found in any of the 4 strains that showed higher levels of knockdown resistance. Little correlation was observed between the presence of thekdrmutation and the level of knockdown resistance, suggesting the existence of multiple resistance mechanisms in many of these strains.  相似文献   

11.
Knockdown resistance (kdr) is a target-site resistance mechanism that confers nerve insensitivity to DDT and pyrethroid insecticides. In the housefly, Musca domestica, molecular cloning of the para-type sodium channel gene has revealed two amino acid mutations that are associated with kdr and super-kdr resistance phenotypes. Both mutations are located in the domain II region of the channel; Leu1014 to Phe in the hydrophobic segment IIS6 and Met918 to Thr in the IIS4-IIS5 linker. To investigate whether these mutations also occur in other insects, we have designed degenerate primers based on conserved sequences in the domain II region of the sodium channel and used these to PCR amplify this region from insecticide-susceptible strains of eight diverse insect species representing four different insect Orders: Helicoverpa armigera, Plutella xylostella, Spodoptera littoralis (Lepidoptera), Blattella germanica (Dictyoptera), Tribolium castaneum (Coleoptera), Myzus persicae, Aphis gossypii and Phorodon humuli (Hemiptera). The primers amplified closely related para-type sodium channel sequences from each insect with a minimum of 85% amino acid identity between species. All of the sequences contained ‘susceptible’ Leu and Met residues at the positions associated with kdr and super-kdr resistance in the housefly. Recent results detailing the presence of a kdr-type Leu to Phe mutation in pyrethroid-resistant strains of two important agricultural pests, P. xylostella and M. persicae, are discussed. ©1997 SCI  相似文献   

12.
RNA-seq data analysis of cigarette beetle (Lasioderma serricorne) strains having different sensitivities to pyrethroids identified sodium channel mutations in strains showing pyrethroid resistance: the T929I and F1534S mutations. These results suggest that reduced sensitivity of the sodium channel confers the pyrethroid resistance of L. serricorne. Results also showed that the F1534S mutation mostly occurred concurrently with the T929I mutation. The functional relation between both mutations for pyrethroid resistance is discussed.  相似文献   

13.
Head lice from Florida (SF-HL) and California (SC-HL) were resistant to permethrin compared with colonized susceptible lice from Panama (PA-HL) (5.5- and 3.4-fold, respectively) and Ecuador (EC-HL) (8.5- and 5.3-fold, respectively). Permethrin-resistant lice were cross-resistant to pyrethrum and DDT. DNA sequencing validated presence of kdr-type mutations (T929I and L932F). Permethrin resistance was synergized by piperonyl butoxide (PBO) in SC-HL. Resistance to malathion in SF-HL (1.4–2.2-fold) and SC-HL (2.1–3.6-fold) was detected. Malathion resistance in SF-HL was synergized by S,S,S-tributylphosphorotrithioate (DEF) and by PBO in SC-HL. Malathion/permethrin-resistant lice from the UK (BR-HL) were synergized by DEF but not synerziged by PBO. PBO protected BR-HL from malathion, indicating suppressed desulfuration. Abamectin resistance in SF-HL (1.7–2.5-fold) and SC-HL (1.8–2.3-fold) was detected. No resistance to lindane was found. Thus, multiple resistance mechanisms against commonly available and widely used pediculicides and insecticides are apparently occurring.  相似文献   

14.
Sodium channel mutations were investigated through nucleotide sequencing of three cDNA fragments amplified from permethrin resistant and susceptible Aedes aegypti from northern Thailand. There was a novel nucleotide substitution (T → G) at the second position of codon 1552 resulting in the replacement of Phenylalanine by Cysteine in segment 6 domain III. This amino acid was indicated by another study to involve an aromatic-aromatic contact between the sodium channel protein and the first aromatic ring of the pyrethroid alcohol moiety. Reciprocal crosses between the homozygous parental susceptible and resistant strains indicated that resistance was autosomal and incompletely recessive, and highly associated with the homozygous mutation. The bioassay of the F2 progeny, formed by backcrossing the F1 with the resistant parental strain, did not show a clear plateau curve across the range of doses, suggesting that resistance to permethrin was controlled by more than one gene locus. Other possible resistance mechanisms involved are discussed.  相似文献   

15.
Pyrethroid and organophosphate resistance-associated mutations have been recently reported in the whitefly Bemisia tabaci (Gennadius), a major pest of protected and outdoor crops worldwide. Here, we developed simple PCR–agarose gel visualization based assays for reliably monitoring the L925I and T929V pyrethroid resistance mutations in the B. tabaci para-type voltage gated sodium channel and the iAChE F331W organophosphate resistance mutation in the acetylcholinesterase enzyme ace1.PCR-RFLP assays were developed for detecting the L925I and the F331W resistance mutations. A highly specific PASA was developed for detecting the T929V mutation. The molecular diagnostic tools were used to monitor the frequency of the resistance mutations in a large number of field caught Q biotype B. tabaci from Crete (Greece), where both organophosphates and pyrethroids are extensively used. The F331W mutation was fixed in all field individuals examined. The pyrethroid resistance mutations were detected in high frequencies: 0.38 and 0.54 for L925I and T929V, respectively. The simple diagnostics are accurate and robust, to be used alongside classical bioassays to prevent ineffective insecticide applications, and for early identification of the spreading of resistant Q biotype populations into new regions around the globe.  相似文献   

16.
Pollen beetle, Meligethes aeneus F. (Coleoptera: Nitidulidae) is a major univoltine pest of oilseed rape in many European countries. Winter oilseed rape is cultivated on several million hectares in Europe and the continuous use of pyrethroid insecticides to control pollen beetle populations has resulted in high selection pressure and subsequent development of resistance. Resistance to pyrethroid insecticides in this pest is now widespread and the levels of resistance are often sufficient to result in field control failures at recommended application rates. Recently, metabolic resistance mediated by cytochrome P450 monooxygenases was implicated in the resistance of several pollen beetle populations from different European regions. Here, we have also investigated the possible occurrence of a target-site mechanism caused by modification of the pollen beetle para-type voltage-gated sodium channel gene. We detected a single nucleotide change that results in an amino acid substitution (L1014F) within the domain IIS6 region of the channel protein. The L1014F mutation, often termed kdr, has been found in several other insect pests and is known to confer moderate levels of resistance to pyrethroids. We developed a pyrosequencing-based diagnostic assay that can detect the L1014F mutation in individual beetles and tested more than 350 populations collected between 2006 and 2010 in 13 European countries. In the majority of populations tested the mutation was absent, and only samples from two countries, Denmark and Sweden, contained pollen beetles heterozygous or homozygous for the L1014F mutation. The mutation was first detected in a sample from Denmark collected in 2007 after reports of field failure using tau-fluvalinate, and has since been detected in 7 out of 11 samples from Denmark and 25 of 33 samples from Sweden. No super-kdr mutations (e.g. M918T) known to cause resistance to pyrethroids were detected. The implications of these results for resistance management strategies of pollen beetle populations in oilseed rape crops are discussed.  相似文献   

17.
Populations of the codling moth, Cydia pomonella L (Lepidoptera, Tortricidae) have developed resistance to several classes of insecticide such as benzoylureas, juvenile hormone analogues, ecdysone agonists and pyrethroids, but the corresponding resistance mechanisms have not been extensively studied. Knockdown resistance (kdr) to pyrethroid insecticides has been associated with point mutations in the para sodium channel gene in a great variety of insect pest species. We have studied two susceptible strains (S and Sv) and two resistant strains (Rt and Rv) of C pomonella that exhibited 4- and 80-fold resistance ratios to deltamethrin, respectively. The region of the voltage-dependent sodium channel gene which includes the position where kdr and super-kdr mutations have been found in Musca domestica L was amplified. The kdr mutation, a leucine-to-phenylalanine replacement at position 1014, was found only in the Rv strain. In contrast, the super-kdr mutation, a methionine-to-threonine replacement at position 918, was not detected in any C pomonella strain. These data allowed us to develop a PCR-based diagnostic test (PASA) to monitor the frequency of the kdr mutation in natural populations of C pomonella in order to define appropriate insecticide treatments in orchards.  相似文献   

18.
为明确河南省部分地区的多花黑麦草Lolium multiflorum种群对乙酰辅酶A羧化酶（acetylCoA carboxylase,ACCase）和乙酰乳酸合成酶（acetolactate synthase,ALS）抑制剂类除草剂的抗性水平和抗性机理,采用整株生物测定法测定采自新乡市和驻马店市的多花黑麦草种群对ACCase抑制剂类除草剂精噁唑禾草灵、炔草酯、唑啉草酯和ALS抑制剂类除草剂甲基二磺隆、氟唑磺隆、啶磺草胺的抗性水平,并对多花黑麦草ACCase和ALS靶标酶编码基因进行克隆及氨基酸序列比对,分析其靶标抗性机理。结果显示,与多花黑麦草敏感种群HNXX01相比,HNZMD04和HNXX05种群对6种除草剂均产生了抗性,HNZMD04种群对精噁唑禾草灵和啶磺草胺的相对抗性倍数分别为44.65和40.31,对炔草酯和氟唑磺隆的相对抗性倍数分别为11.91和11.93;HNXX05种群对精噁唑禾草灵和氟唑磺隆的相对抗性倍数分别为27.70和25.67。HNZMD04和HNXX05抗性种群的ACCase基因均发生了D2078G突变,2个种群的突变率分别为55%和70%;HNZMD04...  相似文献   

19.
为明确天津市小麦田荠菜种群对双氟磺草胺的抗性情况及可能的抗性机理,本研究在天津市静海区、武清区、宝坻区及蓟州区等荠菜发生严重地区的小麦田共采集到6个荠菜种群,采用整株水平测定法测定了6个荠菜种群对双氟磺草胺的抗性水平,并扩增、比对了其靶标乙酰乳酸合成酶 (ALS) 基因部分片段的差异。结果表明:6个荠菜种群对双氟磺草胺均产生了高抗性,抗性倍数在11.4~47.2之间。对抗性和敏感种群的ALS基因片段进行测定比对发现,6个荠菜种群ALS基因197位氨基酸均由脯氨酸 (CCT) 突变为丝氨酸 (TCT),该突变可能是导致荠菜种群对双氟磺草胺产生抗性的重要原因之一。  相似文献   

20.
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  相似文献   

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