几种代谢酶基因与柑橘全爪螨对双甲脒抗性的关系

为了明确羧酸酯酶(carboxylesterase,CarE)基因、谷胱甘肽S-转移酶(glutathione S-transferases,GST)基因和过氧化氢酶(catalase,CAT)基因与柑橘全爪螨Panonychus citri对双甲脒抗性的关系,通过BLAST检索,从柑橘全爪螨转录组数据库中对这3种代谢酶抗性相关基因进行鉴定,并采用RPKM法对双甲脒抗性品系和敏感品系代谢抗性相关基因进行表达差异分析,对差异较大的基因作定量PCR检测.基因差异性分析发现,抗性品系中有9条CarE基因、12条GST基因及6条CAT基因表达量发生上调,13条CarE基因、12条GST基因和3条CAT基因表达量发生下调;Pc29773nrt、Pcl7807nlg和Unigene31477为上调倍数最高的3个基因,其log2 ratio (RS/SS)分别为12.95、10.81、10.01.定量分析显示,Pc29773 nrt、Pcl7807nlg和Unigene31477的上调倍数分别为3.72、2.03和3.09,Pc29773 nrt和Unigene31477上调显著.研究表明柑橘全爪螨Pc29773nrt和Unigene31477上调与其对双甲脒的抗性相关.     

Effects of permethrin and amitraz on gas exchange and water loss in unfed adult females of Amblyomma americanum (Acari: Ixodidae)

Effects of two acaricides, permethrin and amitraz, with different modes of action, on gas exchange and water balance of the lone star tick, Amblyomma americanum, were examined using a flow-through carbon dioxide (CO₂) and water vapor analyzer. Untreated adult female ticks exhibited a distinct discontinuous gas exchange pattern (DGEP) with no measurable water loss. Similarly, ticks with acetone (a solvent) showed little change in gas exchange and water loss. Topical treatment of ticks with a lethal dose of permethrin caused immediate major water loss and long-lasting high frequency DGEP, with increased amplitude of CO₂ release. A small water loss spike was observed for each CO₂ release. Treatment of ticks with amitraz abolished the DGEP and elicited a long-lasting continuous gas exchange pattern, indicating significantly increased tick metabolism. No detectable water loss was observed during amitraz-elicited continuous gas exchange. Treatment of ticks with a mixture of sub-lethal doses of permethrin and amitraz caused a similar initial major water loss, as in permethrin-treated ticks, followed by a second period of water loss activity at 10–14 h after treatment. Mixture of the two acaricides caused immediate onset of major chaotic CO₂ release activities with a significantly elevated baseline CO₂ emission level. The standard metabolism rate (SMR, VCO₂) of acetone-treated ticks (0.452 μl h⁻¹) was similar to that of the untreated ticks (0.461 μl h⁻¹). Compared to untreated control ticks, permethrin-, amitraz-, and the acaricide mixture-treated ticks exhibited significantly increased SMR (1.054, 1.392 and 1.520 μl h⁻¹ respectively (P < 0.05). Results obtained from this study clearly demonstrated the detrimental effects of permethrin and amitraz on CO₂ gas exchange and water balance in A. americanum ticks. The data also revealed subtle differences between permethrin and amitraz in their effects on tick metabolism, specifically gas exchange and water loss. In addition, the study may provide insight into the modes of action of these two acaricides in tick physiology. The results may also help understand the mechanism of synergism between permethrin and amitraz that was reported previously in other ticks and insects.     

Hydrolysis of permethrin by pyrethroid esterases from resistant and susceptible strains of Amblyseius fallacis (Acari: Phytoseiidae)

Pyrethroid-hydrolyzing activity in whole body homogenates of three insecticide-resistant and one susceptible strain of the predator mite, Amblyseius fallacis Garman has been examined in vitro. The highest esterase activity was found in the synthetic pyrethroid-resistant GH-1 strain body homogenate. All three pyrethroid-resistant strains had esterases that hydrolyzed trans-permethrin two times faster than cis-permethrin but isomer specificity was not observed in the susceptible strain. The pyrethroid esterases of the GH-1 strain were very sensitive to inhibition by dichlorovos, S,S,S-tributhylphosphorotrithioate, and 3-octylthio-1,1,1-trifluoro-2-propanone. Carbaryl and tetraethylpyrophosphate exhibited moderate inhibition in the GH-1 strain. Eserine sulfate and piperonyl butoxide only inhibited permethrin hydrolysis at higher concentrations. Fifteen esterase bands were resolved from body homogenates by gradient gel electrophoresis in the GH-1 strain, and were identified as carboxylesterases. The major pyrethroid-hydrolyzing activity was located in E5–E12 bands from GH-1 and composite susceptible strain esterases. Six esterase bands exhibiting low pyrethroid-hydroloyzing activity were also obtained from the two spotted spider mite, Tetranychus urticae (Koch).     

Effect of imidacloprid on the reproduction of acaricide-resistant and susceptible strains of Tetranychus urticae Koch (Acari: Tetranychidae)

Occasional reports linking neonicotinoid insecticide applications to field population outbreaks of the two-spotted spider mite, Tetranychus urticae Koch, have been a topic of concern for integrated pest management (IPM) programmes, particularly in apples. In order to shed light on the factors which may contribute to the occasional field population increase of T. urticae following the application of neonicotinoid insecticides, greenhouse experiments have been set up. Four different T. urticae strains, namely GSS (acaricide-susceptible), WI (organophosphate-selected), USA (a largely uncharacterised strain) and Akita (METI (mitochondrial electron transport inhibitor) acaricide-resistant and cross-resistant to dicofol), were compared for their fecundity without insecticide treatment and for their ovipositional response to foliar and drench applications of the field-relevant dose of imidacloprid (100 mg litre(-1)). Without insecticide treatment, strain GSS laid significantly more eggs (162.50 (+/- 5.43)) than the multiple resistant strain Akita (139.90 (+/- 5.54)) during a 16 day oviposition period. With imidacloprid treatment the highest effect was observed with GSS, with a significantly reduced number of eggs in drench (143.40 (+/- 4.22)) and foliar (144.60 (+/- 5.85)) applications. For strains Akita and USA, no significant differences were observed in oviposition between imidacloprid treatments and controls. The proportion of F1 female offspring decreased significantly with drench application for GSS and WI, while no differences were observed among strains in the survival of F1 immature stages, except for strain USA. The viability of eggs was relatively high (from 82.9 (+/- 4.5)% for USA to 95.2 (+/- 1.2)% for GSS) and not affected by imidacloprid treatments.     

Acaricide toxicity and resistance in larvae of different strains of Tetranychus urticae and Panonychus ulmi (Acari: Tetranychidae)

The toxicities of eight structurally different acaricidal compounds to six‐legged larvae (first motile stage) of three laboratory strains of the two‐spotted spider mite, Tetranychus urticae, and the European red mite, Panonychus ulmi, were evaluated following spray application. The larvae of five field‐derived strains of T urticae originating from France, Italy, Brazil, California and Florida were also tested for their susceptibilities to discriminating concentrations of several acaricides resulting in 95% mortality when applied to the organophosphate‐resistant laboratory reference strain WI. The spray bioassay used was robust and gave repeatable results with a wide range of acaricidal compounds, irrespective of their mode of action (ovo‐larvicides or primarily acting on motile life stages). Compounds tested were abamectin, azocyclotin, chlorpyrifos, clofentezine, deltamethrin, fenpyroximate, hexythiazox and pyridaben. Larvae of one of the laboratory strains of T urticae, AK, originally collected in Japan in 1996 and maintained without further selection pressure, exhibited 2000‐ and >4000‐fold resistance to the mitochondrial electron transport inhibitors pyridaben and fenpyroximate, respectively. Another strain of T urticae, AU, obtained from Australia and maintained in the laboratory under selection with hexythiazox and clofentezine since 1987 showed >770‐ and >1000‐fold resistance to clofentezine and hexythiazox, respectively. The same resistance pattern was observed against larvae of a laboratory strain of P ulmi, CE, also selected with hexythiazox. Larvae of one of the field‐derived strains of T urticae, BR, showed a lower susceptibility to a number of compounds, whilst the others were susceptible to all compounds except the organophosphates. © 2001 Society of Chemical Industry     

Neurophysiological studies of the effects of permethrin upon pyrethroid resistant (kdr) and susceptible strains of dipteran larvae

Permethrin at concentrations as high as 10^?6 had no detectable effect upon neuromuscular transmission at 20-21°C in the body-wall muscles of susceptible larvae of Lucilia sericata and susceptible and resistant (kdr) strains of larvae of Musca domestica. At the higher concentrations of 5 × 10^?6M, muscle cells in all strains and species of larva were slowly depolarised, resulting in failure of neuromuscular transmission when the level of depolarisation corresponded with the peak height of the postsynaptic potentials. Multiple postsynaptic potentials were sometimes obtained in response to a single electrical stimulus to the motor axon. Neither of these effects is considered to result from interference with release of the neurotransmitter or its binding to the postsynaptic receptors. Spontaneous activity in sensory nerve fibres from susceptible larvae was increased followed by intermittent bursting and block at concentrations of permethrin as low as 10^?9M at 20-21°C and 10^?10 at 26°C. Resistant larvae of M. domestica were 1000 times less sensitive, needing concentrations of permethrin as high as 10^?6M at 20-21°C and 10^?7M at 26°C to affect sensory discharge. The results are discussed in relationship to the cause of knockdown by pyrethroid insecticides and to possible changes in the nervous system which may be brought about by the kdr resistance factor.     

Incidence and inheritance of resistance to METI-acaricides in European strains of the two-spotted spider mite (Tetranychus urticae) (Acari: Tetranychidae)

A strain of Tetranychus urticae (Koch; Acari: Tetranychidae), collected from hops (Humulus humuli L; Cannabaceae) in England with a short history of tebufenpyrad use, exhibited resistance to four METI (mitochondrial electron transport inhibitor)-acaricides; tebufenpyrad, pyridaben, fenazaquin and fenpyroximate. Resistance factors for these compounds in a microimmersion assay were 46, 346, 168 and 77 respectively, and corresponded to those exhibited by a Japanese METI-acaricide-resistant reference strain. Levels of resistance remained stable without further selection, and selection with tebufenpyrad did not increase them. The UK strain was also resistant (c 6-fold) to bifenthrin. Crosses of homozygous, diploid females with hemizygous, haploid males showed that, in the UK strain, METI-acaricide resistance was paternally and maternally inherited, and was an incompletely dominant trait. Another tebufenpyrad-resistant strain from the UK, originating from a chrysanthemum nursery (Chrysanthemum foeniculaceum Giseke; Asteraceae) was collected eight months later at a site c 210 km distant from the first. These are the first published incidences of METI-acaricide resistance in Europe and implications for the future use of these compounds are discussed.     

新型杀螨剂-乙唑螨腈对草莓二斑叶螨防效及安全性研究

以二斑叶螨为Tetranychus urticae Koch试虫,采用田间药效试验的方法,研究了乙唑螨腈等6种杀螨剂对二斑叶螨的田间防效及对草莓的安全性。以期为草莓二斑叶螨防控提供高效、安全的杀螨剂。结果表明:30%乙唑螨腈悬浮剂速效性与持效性最好,药后1d的防效为91.80%,14d防效为95.56%;20%丁氟螨酯悬浮剂和100g/L联苯菊酯乳油2种药剂的速效性较好,持效性较差,药后Id防效在83%-89%,药后14d的防效在27%~46%;43%联苯肼酯悬浮剂在药后3d的防效最高,达到87.87%;1.8%阿维菌素乳油在药后1~14d防效在36%-72%之间。73%炔螨特乳油在365mg/L浓度下对草莓叶片产生药害,其它药剂对草莓安全。     

Toxicological and biochemical characterization of coumaphos resistance in the San Roman strain of Boophilus microplus (Acari: Ixodidae)

The San Roman strain of the southern cattle tick, Boophilus microplus, collected from Mexico was previously reported to have a high level of resistance to the organophosphate acaricide coumaphos. An oxidative detoxification mechanism was suspected to contribute to coumaphos resistance in this tick strain, as coumaphos bioassay with piperonyl butoxide (PBO) on larvae of this resistant strain resulted in enhanced coumaphos toxicity, while coumaphos assays with PBO resulted in reduced toxicity of coumaphos in a susceptible reference strain. In this study, we further analyzed the mechanism of oxidative metabolic detoxification with synergist bioassays of coroxon, the toxic metabolite of coumaphos, and the mechanism of target-site insensitivity with acetylcholinesterase (AChE) inhibition kinetics assays. Bioassays of coroxon with PBO resulted in synergism of coroxon toxicity in both the San Roman and the susceptible reference strains. The synergism ratio of PBO on coroxon in the resistant strain was 4.5 times that of the susceptible strain. The results suggested that the cytP450-based metabolic detoxification existed in both resistant and susceptible strains, but its activity was significantly enhanced in the resistant strain. Comparisons of AChE activity and inhibition kinetics by coroxon in both susceptible and resistant strains revealed that the resistant San Roman strain had an insensitive AChE, with a reduced phosphorylation rate, resulting in a reduced bimolecular reaction constant. These data indicate a mechanism of coumaphos resistance in the San Roman strain that involves both insensitive AChE and enhanced cytP450-based metabolic detoxification.     

Acaricidal activity of Cnidium officinale rhizome-derived butylidenephthalide against Tyrophagus putrescentiae (Acari: Acaridae)

The acaricidal activity of materials derived from the rhizome of Cnidium officinale Makino against Tyrophagus putrescentiae Schrank adults was examined using direct contact application and fumigation methods, and compared with that of benzyl benzoate and N,N-diethyl-m-toluamide (DEET). The biologically active constituent of the Cnidium rhizome was identified as butylidenephthalide by spectroscopic analyses. On the basis of 24-h LD50 values, acaricidal activity was more pronounced with butylidenephthalide (5.80 microg cm(-2)) than with benzyl benzoate (9.75 microg cm(-2)) and DEET (16.26 microg cm(-2)). In typical poisoning symptoms, butylidenephthalide resulted in lethargy of treated mites, leading to death without knockdown, whereas benzyl benzoate and DEET caused death associated with uncoordinated behaviour. In a fumigation test with T. putrescentiae adults, butylidenephthalide was much more effective in closed containers than in open ones, indicating that the effects of this compound were largely due to action in the vapour phase. Naturally occurring C. officinale rhizome-derived materials merit further study as potential T. putrescentiae control agents or as lead compounds.     

Characterization of acetylcholinesterases of acaricide resistant and susceptible strains of the cattle tick Boophilus microplus (Can.): II. The substrate specificity and catalytic efficiency of the critical enzyme component

Concentrations and turnover numbers have been determined for the critical acetylcholinesterase component of the organophosphorus susceptible Yeerongpilly and resistant Biarra and Ridgelands strains. The lower specific activity for the substrate acetylthiocholine in the resistant strains was shown to be due to decreased catalytic efficiency and not to any change in the titer of the enzyme. Thioester analogs of acetylthiocholine, with additional methylene groups between the carbonyl carbon and the quaternary nitrogen of the molecule and in the alkyl moiety, have been used to derive K_m and V_max values for the enzymes from each strain. An hypothesis is proposed concerning the alteration of the active site that has occurred to produce the dissimilar aberrant enzymes of the resistant strains and the implications of this hypothesis for inhibitor design are discussed.     

Thermal behaviour and biological activity against Aedes aegypti (Diptera: Culicidae) of permethrin and pyriproxyfen in a smoke‐generating formulation

BACKGROUND: The most common ways to control dengue vector Aedes aegypti (L.) are larval source reduction in domestic habitats and ground application of small quantities of aerosol insecticide (ultralow volume). Nevertheless, these actions have been shown repeatedly to be ineffective in controlling Ae. aegypti populations. RESULTS: The efficacy of a new smoke‐generating formulation containing pyriproxyfen and permethrin was evaluated in the laboratory. Smoke‐generating tablets containing each insecticide individually or combined were prepared, and the recovery of the insecticides from the smoke was determined. Recovery values of over 90% were obtained for pyriproxyfen, and around 50% for permethrin. The biological efficacy of pyriproxyfen released in the smoke was evaluated in the laboratory, on late third‐instar or early fourth‐instar Ae. aegypti larvae, using different concentrations of pyriproxyfen and exposure times. Adult emergence inhibition (EI) values of 100% were obtained at 30 min, and a dose‐dependent effect was observed at 5 min. The effect of pyriproxyfen released in the smoke was due to direct contact with the larvicide in the water rather than by inhalation of the fumes. The efficacy of permethrin released in the fumes was also evaluated as knockdown effect (KT₅₀) on adults for a tablet containing permethrin alone or permethrin plus pyriproxyfen. There was no significant difference in KT₅₀ values obtained for permethrin (KT₅₀ = 19.9 min) and permethrin plus pyriproxyfen (KT₅₀ = 19.4 min). CONCLUSION: The excellent laboratory performance of this new formulation on immature stages and adults indicates that a smoke‐generating tablet containing pyriproxyfen and permethrin could be a new tool for controlling mosquitoes. Copyright © 2009 Society of Chemical Industry     

Mutations in the mitochondrial cytochrome b of Tetranychus urticae Koch (Acari: Tetranychidae) confer cross‐resistance between bifenazate and acequinocyl

BACKGROUND: Resistance of Tetranychus urticae Koch to bifenazate was recently linked with mutations in the mitochondrial cytochrome b Q_o pocket, suggesting that bifenazate acts as a Q_o inhibitor (Q_oI). Since these mutations might cause cross‐resistance to the known acaricidal Q_oI acequinocyl and fluacrypyrim, resistance levels and inheritance patterns were investigated in several bifenazate‐susceptible and bifenazate‐resistant strains with different mutations in the cd1 and ef helices aligning the Q_o pocket. RESULTS: Cross‐resistance to acequinocyl in two bifenazate‐resistant strains was shown to be maternally inherited and caused by the combination of two specific mutations in the cytochrome b Q_o pocket. Although most investigated strains were resistant to fluacrypyrim, resistance was not inherited maternally, but as a monogenic autosomal highly dominant trait. As a consequence, there was no correlation between cytochrome b genotype and fluacrypyrim resistance. CONCLUSIONS: Although there is no absolute cross‐resistance between bifenazate, acequinocyl and fluacrypyrim, some bifenazate resistance mutations confer cross‐resistance to acequinocyl. In the light of resistance development and management, high prudence is called for when alternating bifenazate and acequinocyl in the same crop. Maternally inherited cross‐resistance between bifenazate and acequinocyl reinforces the likelihood of bifenazate acting as a mitochondrial complex III inhibitor at the Q_o site. Copyright © 2009 Society of Chemical Industry     

Phenobarbital induction of permethrin detoxification and phenobarbital metabolism in susceptible and resistant strains of the beet armyworm Spodoptera exigua (Hübner)

The permethrin resistant strain (TR-strain) of the beet armyworm, Spodoptera exigua (Hübner), has 92.5-fold resistance to permethrin (at LD₅₀ level) compared to the permethrin susceptible strain (TS-strain). Bioassay involving permethrin mixed with piperonyl butoxide, an inhibitor of microsomal cytochrome P450s, significantly reduced the resistance ratio from 92.5- to 7.9-fold. However, S,S,S-tributylphosphorotrithioate and diethylmaleate which are inhibitors of esterases and glutathione S-transferase, respectively, did not affect the resistance level. These results indicate that the detoxification of permethrin in the TR-strain was primarily due to the cytochrome P450 monooxygenases. LD₅₀ for permethrin was increased to 4.5-fold by the pre-treatment of phenobarbital in the TS-strain. The effect of induction by phenobarbital was almost completely overcome by the piperonyl butoxide treatment. However, it was observed that phenobarbital treatment did not cause any change in the toxicity of permethrin to TR strain. Since this result deviated from the expectation that the metabolism of phenobarbital in the TR-strain should be greater than that in the TS-strain, it was deemed necessary to compare the metabolism of phenobarbital between the TS- and TR-strains. Comparison was made based on the concentration of phenobarbital in the hemolymph and whole body. The results showed no significant difference in phenobarbital treatment between the two strains used in this study suggesting the possibility that the induction system in TS-strain is different from the TR-strain.     

Acaricidal activities of paeonol and benzoic acid from Paeonia suffruticosa root bark and monoterpenoids against Tyrophagus putrescentiae (Acari: Acaridae)

The acaricidal activities of paeonol (2'-hydroxy-4'-methoxyacetophenone) and benzoic acid identified in the root bark of tree peony, Paeonia suffruticosa Andrews, against copra mite, Tyrophagus putrescentiae (Schrank), adults were examined using direct contact and vapour phase toxicity bioassays and compared with those of cinnamyl acetate, cinnamyl alcohol and 37 monoterpenoids as well as the acaricides benzyl benzoate, dibutyl phthalate and N,N-diethyl-m-toluamide (DEET). Based on LD(50) values in fabric piece contact toxicity bioassays, the acaricidal activities of benzoic acid (4.80 microg cm(-2)) and paeonol (5.29 microg cm(-2)) were comparable to that of benzyl benzoate (4.46 microg cm(-2)) but more pronounced than those of DEET (30.03 microg cm(-2)) and dibutyl phthalate (25.23 microg cm(-2)). In vapour phase toxicity bioassays, paeonol and benzoic acid were much more effective in closed containers than in open ones, indicating that the effects of these compounds were largely due to action in the vapour phase. As judged by 24 h LD(50) values, (1S)-(-)-verbenone (7.42 mg per disc) was the most toxic fumigant, followed by (1S)-(-)-camphor, (S)-(+)-carvone, (R)-(-)-linalool and (+/-)-camphor (10.45-18.18 mg). Potent fumigant toxicity was also observed with paeonol, (2S,5R)-(-)-menthone, (+/-)-citronellal, benzoic acid, (1S,4R)-(-)-alpha-thujone and (R)-(+)-pulegone (25.10-34.63 mg). Neither benzyl benzoate, DEET nor dibutyl phthalate caused fumigant toxicity. Paeonia root bark-derived materials, particularly paeonol and benzoic acid, as well as the monoterpenoids described, merit further study as potential acaricides or as leads for the control of T. putrescentiae.     

Insecticide resistance management of permethrin and acephate against the cowpea curculio,Chalcodermus aeneus Boheman (Coleoptera: Curculionidae), a pest of the southern pea,Vigna unguiculata (L.) Walp.

To evaluate strategies for management of potential resistance of the cowpea curculio, Chalcodermus aeneus Boheman, to sprayer‐applied permethrin and acephate, five management regimes were evaluated on early, midseason and late sequential plantings of southern peas, Vigna unguiculata (L.) Walp. during 1991–1993. Management regimes were: (1) consecutive permethrin (0.11 kg a.i./ha early‐midseason‐late); (2) consecutive acephate (1.1 kg a.i.ha early‐midseason‐late); (3) consecutive mixture (0.055kg a.i./ha permethrin and 0.55kg a.i./ha acephate early‐midseason‐late); (4) alternation (0.11 kg a.i./ha permethrin early and 1.1 kg a.i./ha acephate midseason and late plantings); and (5) an untreated check early, midseason and late. The effect of management regime on the toxicities (LC₅₀) of permethrin and acephate was determined by topical application to next generation adult cowpea curculios reared from infested southern pea pods harvested from untreated plants in the management fields. In the field, infestation varied more by planting date than by management regime. Toxicity of permethrin to adults selected in the consecutive mixture regime was significantly lower in 1991 and 1992 late season and in 1993 midseason than in early season for the respective years. There was a seasonal decrease in the laboratory toxicity of acephate to adults with all management regimes.