The biochemical basis of resistance to organophosphorus insecticides in the sheep blowfly, Lucilia cuprina

The metabolism in vivo and in vitro of [¹⁴C]parathion and [¹⁴C]paraoxon was studied in a susceptible (LS) and an organophosphorus-resistant (Q) strain of the sheep blowfly, Lucilia cuprina. Both strains detoxified the insecticides in vivo via a number of pathways, but the resistant strain produced more of the metabolites diethyl phosphate and diethyl phosphorothionate. No difference was found between strains in the rate of penetration of the compounds used. Also, in vitro studies showed no difference between strains in the sensitivity of head acetylcholinesterase to inhibition by paraoxon. Both the microsomal and the 100,000g supernatant fractions degraded paraoxon, but resistance in Q could be explained by the eightfold greater rate of diethyl phosphate production with or without added NADPH. Parathion was also degraded to diethyl phosphorothionate by an NADPH-requiring enzyme in microsomal preparations from both strains. However, Q produced significantly more diethyl phosphorothionate in vivo than LS. It was concluded that organophosphorus resistance in Q was due mainly to a microsomal phosphatase hydrolyzing phosphate but not phosphorothionate esters, probably enhanced by a microsomal oxidase detoxifying the latter.     

Cross-resistance to dicyclanil in cyromazine-resistant mutants of Drosophila melanogaster and Lucilia cuprina

Cyromazine is an insect growth regulator insecticide with a novel mode of action, mainly used to control dipteran insects. Previously, cyromazine-resistant mutants of the Australian sheep blowfly Lucilia cuprina and the vinegar fly Drosophila melanogaster have been isolated following ethyl methanesulfonate mutagenesis and selection for resistance. Here, we show that these cyromazime-resistant mutants are cross-resistant to dicyclanil, an insect growth regulator compound with a similar chemical structure to cyromazine. Dicyclanil was recently introduced as a control agent of L. cuprina. Cross-resistance to the benzylphenylurea insecticide lufenuron was also assessed. Only one D. melanogaster cyromazine-resistant mutant is cross-resistant to lufenuron.     

Arginine kinase of the sheep blowfly Lucilia cuprina: Gene identification and characterization of the native and recombinant enzyme

Arginine kinase (ATP:l-arginine ω-N-phosphotransferase, EC2.7.3.3.; AK) is an enzyme with crucial functions in the energy metabolism of insects and other invertebrates as well as some protozoa and has been proposed as a parasiticidal and pesticidal drug target. In this study we report the identification, cDNA cloning, genomic gene structure and functional expression of an AK gene from the parasitic sheep blowfly Lucilia (L.) cuprina. The blowfly AK-encoding gene is devoid of introns and present in a single copy in the L. cuprina genome. AK is present in L. cuprina flies as a highly expressed soluble enzyme and is more abundant in the thorax, where it represents up to 2% of the total soluble protein, compared to head or abdomen. Guanidino substrate specificity studies show that L. cuprina AK is stereospecific for the l-form over the d-form of its specific substrate arginine. Furthermore, the presence of a free or esterified carboxylic group as well as an unsubstituted α-amine group are essential for acceptance of the l-arginine substrate while modifications in the aliphatic side chain are better tolerated. The apparent Michaelis-Menten constants K_M and the molecular size of the recombinant enzyme are virtually identical to that of the native enzyme suggesting that the gene cloned in this study encodes the highly expressed AK enzyme of the sheep blowfly.     

Synergism of DDT effects by DMC on labellar receptors of resistant and susceptible house flies

Electrophysiological responses of labellar hairs of resistant and susceptible strains of the house fly were recorded at times following treatment of the hairs with DDT. Under the influence of DDT, the receptors of a hair discharged groups of impulses in high-frequency trains instead of the usual regular volley. The effect was observed to gradually spread to nontreated hairs. Among four strains chosen for gradation from high resistance to high susceptibility, in general the relative effectiveness of DDT corresponded to overall resistance as indicated by LD₅₀ data. The highly resistant strain showed essentially no effects, and the other strains showed effects with some degree of recovery. Results for the highly susceptible strain were anomalous in not differing significantly from those of the moderately resistant strai. Unexpectedly small effects in the highly susceptible strain point to a strain characteristic not necessarily correlated with LD₅₀ data.     

不同药剂对山楂叶螨的亚致死效应

在室内采用叶碟饲养的方法,利用生命表技术研究了四螨嗪、阿维菌素、甲氰菊酯及三唑锡对山楂叶螨的亚致死效应.结果表明,在成虫期,LC10、LC25剂量的阿维菌素、甲氰菊酯及三唑锡可使成螨的产卵量显著降低,除甲氰菊酯LC10外,其余处理可使次代种群的内禀增长率（rm）由0.173降低至0.133～0.167;在若螨Ⅰ期,LC10、LC25剂量的阿维菌素可使rm由0.237降低至0.197～0.199;甲氰菊酯LC10对雌螨的产卵有明显的刺激作用,rm由0.237提高到0.259,而LC25则无明显影响;在卵期,四螨嗪LC10对rm无明显影响,但LC50、LC25可使rm由0.173降低至0.149～0.154.     

五种常用杀虫剂对灰飞虱繁殖力的影响

为合理使用杀虫剂防治灰飞虱,采用稻茎浸渍法测定杀虫剂氟虫腈、吡虫啉、毒死蜱、三唑磷和高效氯氟氰菊酯对灰飞虱3龄若虫的室内活性,并用LC50、LC75和LC,90测定灰飞虱的繁殖力.结果表明,经氟虫腈和吡虫啉处理的3龄若虫和成虫,所产的第2代若虫数均少于清水对照;而经毒死蜱、三唑磷和高效氯氟氰菊酯处理的3龄若虫和成虫,所产的第2代若虫数均多于对照,其中经高效氯氟氰菊酯LC50处理的3龄若虫所产的第2代若虫数最多,比对照多53.3%,极显著高于对照.在尼龙网隔离的水泥池内稻麦套作,接灰飞虱成虫后分别用氟虫腈39.9 mL/667m2和吡虫啉53.2mL/667m2处理,第2年的虫量仅为对照的33.5%和64.6%;而用毒死蜱59.9mL/667m2、三唑磷98.6mL/667 m2以及高效氯氟氰菊酯99.8mL/667m2处理,第2年的虫量分别比对照多1.73、1.35和1.93倍.     

Effects of mutations in Drosophila nicotinic acetylcholine receptor subunits on sensitivity to insecticides targeting nicotinic acetylcholine receptors

Several strains of Drosophila melanogaster possess mutant alleles in nicotinic acetylcholine receptor (nAChR) subunits, Dα1 and Dβ2 that confer resistance to neonicotinoids such as imidacloprid and nitenpyram, and Dα6, that confers resistance to spinosyns. These mutant strains were bioassayed with a selected set of nAChR active insecticides including neonicotinoids, spinosad, and sulfoxaflor, a new sulfoximine insecticide. All of the neonicotinoids examined, except dinotefuran showed reduced insecticidal efficacy on larvae of the Dα1 mutant, suggesting that this subunit may be important in the action of these insecticides. All of the neonicotinoids, including dinotefuran, showed reduced insecticidal efficacy on larvae possessing the Dβ2 mutation. A similar pattern of broad neonicotinoid resistance to that of Dβ2 alone was also observed for larvae with both the mutations (Dα1 + Dβ2). The Dβ2 mutation exhibited a lower level of cross-resistance to sulfoxaflor (<3-fold) than to any of the neonicotinoids (>13-fold). In contrast, there was no cross-resistance for any of the neonicotinoids or sulfoxaflor in adult flies with the Dα6 mutation, which confers high levels of resistance to spinosad. Thus in the D. melanogaster strains studied, target site resistance observed for the neonicotinoids and the spinosyns does not translate directly to resistance towards sulfoxaflor.     

噻虫胺等药剂对韭菜迟眼蕈蚊的致毒效应

为了明确新烟碱类药剂对韭菜迟眼蕈蚊的毒力,采用管测药膜法和药液定量滴加法测定了噻虫胺等6种药剂对韭菜迟眼蕈蚊不同虫态的毒力,并研究了噻虫胺、甲氨基阿维菌素苯甲酸盐和辛硫磷亚致死浓度对其4龄幼虫生长发育和繁殖的影响。结果表明,新烟碱类杀虫剂噻虫胺、吡虫啉和噻虫嗪对成虫的击倒毒力均较高,分别是阿维菌素的5.75、3.86和3.51倍;6种药剂对韭菜迟眼蕈蚊卵的毒力均较低;对2龄和4龄幼虫的毒力,均以噻虫胺最高,LC₅₀分别为0.339 mg/L和1.020 mg/L,分别是阿维菌素的27.00倍和25.23倍。用噻虫胺亚致死剂量处理韭菜迟眼蕈蚊4龄幼虫,其发育历期和蛹期延长,蛹重、化蛹率、成虫羽化率、单雌产卵量和卵孵化率均降低。     

Structure-related effects of pyrethroid insecticides on the lateral-line sense organ and on peripheral nerves of the clawed frog, Xenopus laevis

The effects of seven different pyrethroid insecticides on the lateral-line sense organ and on peripheral nerves of the clawed frog, Xenopus laevis, were investigated by means of electrophysiological methods. The results show that two classes of pyrethroid can be clearly distinguished. (i) Pyrethroids without an α-cyano group (permethrin, cismethrin, and bioresmethrin). These noncyano pyrethroids induce short trains of nerve impulses in the lateral-line sense organ. In peripheral nerve branches they induce a depolarizing afterpotential and repetitive firing. These effects are very similar to those previously reported for allethrin. (ii) Pyrethroids with an α-cyano-3-phenoxybenzyl alcohol (cypermethrin, fenpropathrin, deltamethrin, and fenvalerate). In the lateral-line sense organ these α-cyano pyrethroids induce very long trains of nerve impulses which may last for seconds and may contain hundreds or even thousands of impulses. The α-cyano compounds do not cause repetitive activity in peripheral nerves. Instead they induce a quickly reversible, stimulus frequency-dependent suppression of the action potential. Since the chemical structure of cypermethrin differs from that of permethrin only in the α-cyano group and because all four α-cyano compounds act in a very similar way, it is concluded that the α-cyano substituent is responsible for the large differences in neurotoxic effects. In the lateral-line sense organ the duration of nerve impulse trains induced by the noncyano as well as the α-cyano pyrethroids increases dramatically when the temperature is lowered. Further, in sensory fibers the effects of both classes of pyrethroid on the nerve action potential are more pronounced compared to their effects on motor fibers. It is argued that the different neurotoxic effects reported here originate from a common mechanism of action of pyrethroids, which is a prolongation of the transient increase of sodium permeability of the nerve membrane associated with excitation.It is concluded that the sodium channel in the nerve membrane is the major target site of noncyano and α-cyano pyrethroids.     

新烟碱类等杀虫剂对葱蝇的毒力及其对生长发育和繁殖的影响

采用定量滴加法和浸渍法测定了8种药剂对葱蝇1龄和3龄幼虫的毒力,用噻虫胺和噻虫嗪等新烟碱类杀虫剂的亚致死浓度分别处理葱蝇1龄和3龄幼虫,观察其对葱蝇生长发育和繁殖的影响,并进行了田间药效试验。结果表明,噻虫胺和噻虫嗪对3龄幼虫48 h的LC₅₀分别为14.7839 mg/L和13.3055 mg/L,毒力为毒死蜱的0.41倍和0.50倍。但LC_30-40剂量处理葱蝇3龄幼虫后,第6天存活幼虫体重分别较对照降低35.47%和36.34%,幼虫发育历期延长3.68天和3.98天,化蛹率降低71.91%和78.65%,雌蛹重降低13.36%和14.72%,雄蛹重降低15.97%和18.32%,羽化率降低12.50%和17.86%,单雌产卵量降低47.71%和51.68%。噻虫胺和噻虫嗪处理1龄幼虫,对其生长发育也有显著不利影响。田间药效试验结果显示,1 200 g/hm²噻虫胺和噻虫嗪处理后7天防效分别达98.16％和92.46％,表明这两种药剂对葱蝇防治效果高、推广应用价值大。     

Toxicity and neurophysiological effects of selected insecticides on the mole cricket, Scapteriscus vicinus (Orthoptera: Gryllotalpidae)

Mole crickets (Scapteriscus spp.) are severe subterranean pests of turfgrasses, commonly targeted with neurotoxic insecticides. Ideally insecticides used against mole crickets should induce quick knockdown or mortality to minimize damage caused by their tunneling. However, neurophysiological effects of insecticides on mole crickets are mostly unknown. The aims of this study were to investigate neurophysiological and toxic effects of several insecticides on tawny mole cricket (Scapteriscus vicinus Scudder) adults and nymphs, and potential synergy between pyrethroid and neonicotinoid insecticides. Bifenthrin, fipronil, and the combination of bifenthrin + imidacloprid provided the fastest median mortality when injected. The combination of bifenthrin + imidacloprid elicited faster toxicity than either active ingredient alone. Imidacloprid, bifenthrin, and bifenthrin + imidacloprid caused immediate knockdown, whereas fipronil immobilized mole crickets within 1-2 h. Acephate, bifenthrin, fipronil, imidacloprid, and bifenthrin + imidacloprid caused significant neuroexcitation. Bifenthrin + imidacloprid resulted in greater increases of spontaneous neural activity than the additive effects of imidacloprid and bifenthrin alone. Excitatory compounds acting at sodium and chloride channels (bifenthrin and fipronil) were the most toxic against S. vicinus. Combining a sodium channel toxin (bifenthrin) and a synaptic toxin (imidacloprid) led to greater than additive neurophysiological and toxic effects, which to our knowledge provides the first documented evidence of synergistic neurological “potentiation”.     

In vitro metabolism of insecticides during midgut penetration

A study of the metabolism of ¹⁴C-labeled dieldrin, DDT, malathion, and carbaryl during penetration of the isolated midgut of two insects (Blaberus discoidalis and Manduca sexta) and a section of the intestine of a mammal (Mus musculus) is reported. There was appreciable metabolism of malathion during penetration, including differences in the activation reaction to malaoxon, between insects and mammals. Metabolism was relatively slow during penetration of carbaryl and the chlorinated hydrocarbon insecticides, and little difference in metabolic patterns was noted among the three species. The penetration studies were supported by experiments in which insecticides were incubated with intact and homogenized midgut preparations.     

The action of insecticides on neurosecretory neurons in the stick insect, Carausius morosus

The action of insecticides on the spontaneous electrical activity of neurohemal tissue in the stick insect, Carausius morosus, has been studied using extracellular electrodes. The pyrethroid, permethrin, causes a massive increase in the frequency of the spontaneously generated action potentials at concentrations between 5 × 10^?5 and 5 × 10^?8M. Concentrations as low as 5 × 10^?11M are still effective in producing bursting activity.DDT, at concentrations between 5 × 10^?5M and 5 × 10^?6M, produces an overall increase in activity although the bursting activity is less violent than that shown with permethrin. DDT, 5 × 10^?7M, is ineffective at altering the resting pattern.Carbaryl and coroxon cause a transitory increase in electrical activity at 1 × 10^?4M, but are ineffective at 1 × 10^?5M.It is concluded that insecticides could have a direct effect upon the neurohormonal balance in insects.     

The effect of insecticides on osmotic and lonic balance in the locust Schistocerca gregaria

Schistocerca poisoned with insecticides may lose more than 30% of their body weight and 60% of their hemolymph volume within 24 hr. In spite of this and the impairment of the nervous system, poisoned Schistocerca show a remarkable ability to control the osmotic concentration and ionic composition of the remaining hemolymph. Much of the water loss from poisoned locusts can be attributed to evaporation through the spiracles, due to the progressive failure of the animal to close the spiracular valves. Another significant factor is a massive transport of fluid across the gut wall into the fore and midgut. This fluid may be regurgitated causing a further loss in weight.     

Effects of DDT on labellar receptors in two DDT-resistant and two DDT-susceptible strains of house flies

Electrophysiological responses of labellar hairs of resistant and susceptible strains of the house fly were recorded at times following treatment of the hairs with DDT. Under the influence of DDT, the receptors of a hair discharged groups of impulses in high-frequency trains instead of the usual regular volley. The effect was observed to gradually spread to nontreated hairs. Among four strains chosen for gradation from high resistance to high susceptibility, in general the relative effectiveness of DDT corresponded to overall resistance as indicated by LD₅₀ data. The highly resistant strain showed essentially no effects, and the other strains showed effects with some degree of recovery. Results for the highly susceptible strain were anomalous in not differing significantly from those of the moderately resistant strai. Unexpectedly small effects in the highly susceptible strain point to a strain characteristic not necessarily correlated with LD₅₀ data.     

异色瓢虫对三种杀虫剂的抗性选育

培育天敌抗药性品系,可以促进天敌在害虫防治中的运用。高效氯氰菊酯、吡虫啉、毒死蜱是应用广泛的新型杀虫剂,适用于防治粮食、果树、蔬菜和其它经济作物的害虫。异色瓢虫Harmonia axyridis（Pallas）是多种蚜虫的捕食性天敌,分布广泛。     

Synergistic and antagonistic effects of atrazine on the toxicity of organophosphorodithioate and organophosphorothioate insecticides to Chironomus tentans (Diptera: Chironomidae)

The acute toxicities of two organophosphorodithioate (dimethoate and disulfoton) and two organophosphorothioate (omethoate and demeton-S-methyl) insecticides were evaluated individually and in binary combination with the herbicide atrazine using fourth-instar larvae of the aquatic midge, Chironomus tentans. Atrazine alone up to 1000 μg/L did not show significant toxicity to the midges in a 48-h bioassay. However, atrazine concentrations as low as 1 μg/L in combination with dimethoate at EC₂₅ (concentration to affect 25% of tested midges), 100 μg/L in combination with disulfoton (EC₂₅), and 10 μg/L in combination with demeton-S-methyl (EC₂₅) significantly enhanced the toxicity of each organophosphate insecticide. In contrast, atrazine concentrations of 10 μg/L and above in combination with omethoate (EC₂₅) significantly decreased the toxicity of the insecticide. Biochemical analysis indicated that increased toxicity of dimethoate, disulfoton, and demeton-S-methyl in binary combination with atrazine correlated to the increased inhibition of acetylcholinesterase. Furthermore, cytochrome P450-dependent O-deethylation activity in the midges exposed to atrazine at 1000 μg/L was 1.5-fold higher than that in the control midges. Thus, atrazine appeared to induce cytochrome P450 monooxygenases in the midges. Elevated cytochrome P450 monooxygenase activity may increase the toxicities of dimethoate, disulfoton, and demeton-S-methyl by enhancing the oxidative activation of dimethoate into omethoate, and disulfoton and demeton-S-methyl into their sulfoxide analogs with increased anticholinesterase activity. In contrast, atrazine reduced the toxicity of omethoate possibly by enhancing the oxidative metabolic detoxification since omethoate does not require oxidative activation.     

七种新烟碱类杀虫剂对韭菜迟眼蕈蚊幼虫及蚯蚓的选择毒力

为筛选出高效安全的韭蛆防治药剂,室内采用胃毒触杀联合毒力法比较了吡虫啉、啶虫脒、噻虫嗪、噻虫胺、呋虫胺、烯啶虫胺、噻虫啉与毒死蜱和高效氯氟氰菊酯等6种对照药剂对韭菜迟眼蕈蚊幼虫的毒力,同时用人工土壤法测定了13种药剂对蚯蚓的急性毒性,并通过盆栽试验验证了其对韭蛆和蚯蚓的选择毒力。结果表明,吡虫啉、噻虫胺、呋虫胺、噻虫啉、噻虫嗪对韭菜迟眼蕈蚊4龄幼虫的毒力明显高于6种对照药剂,对虫酰肼的相对毒力倍数分别为101.6、55.0、32.9、27.2、13.6;13种供试药剂中,除吡虫啉、啶虫脒、噻虫胺、呋虫胺对蚯蚓中等毒性外,其余均为低毒;盆栽试验中,吡虫啉、噻虫嗪、毒死蜱、噻唑膦、高效氯氟氰菊酯的防虫效果和保苗效果均分别高于其它药剂,但其中只有噻虫嗪对蚯蚓没有明显致死作用。     

Deletion of Cyp6d4 does not alter toxicity of insecticides to Drosophila melanogaster

Cytochrome P450-dependent monooxygenases are important in the activation and detoxification of numerous insecticides. In this study, a Drosophila melanogaster Cyp6d4 null mutant was used to determine the role of this P450 in insecticide metabolism. This null mutant was generated by imprecise excision of a mobile P element located upstream to the P450 gene Cyp6d4. Comparative analysis between the non-functional mutant and relevant control strains shows that Cyp6d4 does not appear to be involved in the metabolism of chlorfenapyr, cypermethrin, diazinon, imidacloprid, malathion, oxamyl, parathion, or pyrethrum extract, even though these insecticides are known to be activated or detoxified by P450-monooxygenases. No obvious abnormalities in development were seen in the Cyp6d4 null mutant, indicating that Cyp6d4 is not critical for the metabolism of vital endogenous substrates.     

Penetration of insecticides through the foregut of the honeybee (Apis mellifera L.)

The rates of penetration of ¹⁴C-labeled insecticides (parathion, carbaryl, and dieldrin) through the foregut of the honeybee (Apis mellifera L.) were measured in vitro and in vivo. Uptake of the insecticides from the lumen of the foregut into foregut tissue was directly proportional to insecticide lipophilicity, but penetration through the foregut was not. Of the three insecticides studied, parathion appeared to possess the optimal physicochemical characteristics required for penetration. The uptake of carbaryl and release of dieldrin by the foregut tissues may limit their respective penetration rates. Insecticide penetration was found to be inversely proportional to the sucrose concentration in the lumen of the foregut in both in vitro and in vivo studies. The oral toxicity of carbaryl showed a similar dependence on the sucrose concentration of the solution in which the insecticide was fed. The data presented indicate that the honeybee foregut is permeable to lipophilic compounds and strongly suggest that this permeability may contribute substantially to the toxicity of orally ingested insecticides in this insect.