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1.
Acetylcholinesterase (AChE) was purified by affinity chromatography from two populations of the oriental migratory locust, Locusta migratoria manilensis (Meyen), collected from Huanghua and Pingshan Counties, Hebei Province of China. The purification factors and yields were 1661-fold and 19.3%, respectively, for the Huanghua population, and 3897-fold and 39.6% for the Pingshan population. Both the purification factor and yield were significantly lower in the Huanghua population than in the Pingshan population. AChE activity was almost completely inhibited by 10−6 M eserine and BW284C51, but ?5.8% of AChE activity was inhibited by ethopropazine at the same concentration, suggesting that purified AChE from either population was a typical insect AChE. However, AChE purified from the Huanghua population was 62-, 2.0-, and 1.6-fold less sensitive to inhibition by the three organophosphate compounds, chlorpyrifos oxon, demeton-S-methyl, and paraoxon, respectively, than that from the Pingshan population. Significantly lower purification factor and low yield associated with reduced sensitivity of AChE to inhibition by the organophosphates indicated that AChE purified from the Huanghua population was biochemically and pharmacologically different from that of the Pingshan population. Reduced sensitivity of AChE appeared to contribute to organophosphate resistance in the locust from Huanghua County, where insecticides have commonly been used to manage outbreaks of the locust.  相似文献   

2.
The role of esterases in malathion resistance in Culex tarsalis has been investigated. When larvae of a resistant and a sensitive strain were placed in water containing [14C]malathion, malathion penetrated to give initially similar internal levels. With resistant mosquitoes, after 15 min the internal malathion concentration decreased to low levels while the monoacid degradation products accumulated in the larvae and were excreted into the surrounding water, whereas in susceptible larvae the internal malathion level stayed high and was lethal. It is suggested that the decrease in internal malathion and the resulting resistance were caused by an active malathion carboxylesterase in the resistant strain. A specific assay for malathion carboxylesterase with [14C]malathion showed 55 times more activity in resistant than in susceptible larvae, whereas when general esterase activity was assayed with α-naphthyl acetate only 1.7 times the activity was found. Analyses by starch gel electrophoresis showed a peak of malathion carboxylesterase, 60-fold higher from resistant than from susceptible larvae, in a gel zone which did not stain for general esterase activity. General esterases that did not hydrolyze malathion showed different electrophoretic patterns in the two populations, which are likely due to the nonisogenic character of the strains. These results show that use of a specific assay and the demonstration of degradation of malathion in vivo are essential for assessment of the contribution of esterase activity to the malathion-resistant phenotype in mosquito populations.  相似文献   

3.
The levels of susceptibility of populations of the European red mite Panonychus ulmi (Koch) (Acarina: Tetranychidae) collected from apple orchards in the Bursa region of Turkey to the insecticides chlorpyrifos and lambda-cyhalothrin, were determined by a petri leaf disk—Potter spray tower method. When compared with the susceptible population, resistance ratios at the LC50 level ranged from 6.0- to 35.6-fold, and from 0.7- to 5.7-fold for chlorpyrifos and lambda-cyhalothrin, respectively. Kinetic parameters of general esterase activity with α-naphthyl acetate as substrate indicated that an increased activity was present in the resistant populations compared with the susceptible populations. In these strains, 1.5- and 2.2-fold higher Glutathione S-transferase (GST) activity was also detected with the substrate 1-chloro-2,4-dinitrobenzene. General esterase activity gel profiles of these populations were studied by native polyacrylamide gel electrophoresis, but no relationship between resistance ratios and band patterns was detected. The results of this study document a decreased efficacy of chlorpyrifos and lambda-cyhalothrin in field populations of P. ulmi in Turkey, possibly linked to altered activities of esterases and GST.  相似文献   

4.
Resistance in Spodoptera litura (Fabricius) has been attributed to enhanced detoxification of insecticides by increased levels of esterases, oxidases and/or glutathione S-transferases. Enzyme inhibiting insecticide synergists can be employed to counter increased levels of such enzymes in S. litura. Dihydrodillapiole induced synergism of pyrethroid toxicity was examined in the laboratory-reared third instar larval population of S. litura collected in Delhi (susceptible), and Guntur (resistant) region of Andhra Pradesh, India. The Guntur population was found to be 7.04 and 10.19 times resistant to cypermethrin and lambdacyhalothrin, respectively. The activity of cypermethrin, lambdacyhalothrin and profenophos against susceptible and resistance populations of S. litura, was gradually increased when used along with a plant-derived insecticide synergist dihydrodillapiole. The α-naphthyl acetate hydrolysable esterase activity in Delhi population was less as compared to the Guntur population. Resistance associated esterases in Delhi population were inhibited by pre-treatment with dihydrodillapiole. The esterase level in insect was instantly reduced initially, sustained for about 3 h and equilibrated at 4 h post treatment. The esterase activity of Guntur population was increased to 1.28 μmoles/mg/min at 2 h post treatment and subsequently reduced to lower than 0.70 μmoles at 4-12 h post treatment. The variation in esterase activity is suggestive of its homeostatic regulation in test populations. Dihydrodillapiole thus caused significant reduction of resistance in S. litura to cypermethrin, lambda cyhalothrin and profenophos.  相似文献   

5.
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   

6.
A field-collected strain (MR-VL) of the two-spotted spider mite, Tetranychus urticae Koch, exhibited strong resistance to bifenthrin, dicofol and fenbutatin oxide in comparison with a susceptible laboratory strain (LS-VL). The MR-VL strain was screened for cross-resistance with several currently used acaricides. Cross-resistance was detected with clofentezine (RR = 2631), dimethoate (RR = 250), chlorfenapyr (RR = 154), bromopropylate (RR = 25), amitraz (RR = 17), flucycloxuron (RR = 15) and azocyclotin (RR = 7). Abamectin, acequinocyl, bifenazate, tebufenpyrad and spirodiclofen did not show any signs of cross-resistance. Enhanced detoxification by increased activity of mono-oxygenases (MO) and esterases is at least partially responsible for the observed resistance and cross-resistance. MO assays with 7-ethoxycoumarin (7-EC) were optimised and 7-ethoxy-4-trifluoromethylcoumarin (7-EFC), a new MO-substrate, was evaluated for the first time in T urticae and proved to be a good alternative to 7-EC. Approximately 3- and 4-fold higher MO activity was detected with 7-EFC and 7-EC respectively in the MR-VL strain. Kinetic parameters of general esterase assays with 4-nitrophenyl acetate and 1-naphthyl acetate as substrate indicated that more esterases were present in the MR-VL strain. A first attempt was made to classify the esterases present in T urticae. Acetyl-, aryl- and carboxyl-esterases were detected with the use of inhibitors after separation by native PAGE. Glutathione-S-transferases did not seem to play any role in the observed resistance and no differences were detected when the general oxidative capacities of the two strains were compared.  相似文献   

7.
A series of 27 substituted thio-1,1,1-trifluoropropanones was synthesized by reacting the corresponding thiol with 1,1,1-trifluoro-3-bromopropanone. The resulting sulfides were screened as inhibitors of hemolymph juvenile hormone esterase and α-naphthyl acetate esterase activity of the cabbage looper, Trichoplusia ni, electric eel acetylcholinesterase, bovine trypsin, and bovine α-chymotrypsin. The presence of the sulfide bond increased the inhibitory potency on all of the enzymes tested when compared with compounds lacking the sulfide. In general, the compounds proved to be poor inhibitors of chymotrypsin and moderate inhibitors of trypsin. By varying the substituent on the sulfide, good inhibitory activity was obtained on α-naphthyl acetate esterase, acetylcholinesterase, while some of the compounds proved to be extremely powerful inhibitors of juvenile hormone esterase. The most powerful inhibitor tested was 3-octylthio-1,1,1-trifluoro-2-propanone, with an I50 of 2.3 × 10?9M on JH esterase. This compound showed a molar refractivity similar to that of the JH II backbone, was not toxic to T. ni, and was moderately toxic to mice, with a 48-hr LD50 of >750 mg/kg. It effectively delayed pupation when applied to prewandering larvae of T. ni, as expected for a JH esterase inhibitor. Thus, some members of this series are promising for evaluating the role of JH esterase in insect development. The series also indicates that, by varying the substituent on the sulfide moiety, potent “transition-state” inhibitors can be developed for a wide variety of esterases and proteases.  相似文献   

8.
Esterases hydrolyzing α-naphthyl acetate (α-NA), β-naphthyl acetate (β-NA), and p-nitrophenyl acetate (p-NPA) were investigated colorimetrically in larval homogenates of synthetic pyrethroid susceptible (S) and resistant (R) strains of Spodoptera littoralis (Boised). The hydrolytic activity towards the three substrates in cybolt, decamethrin, and fenvalerate R strains were from 3 to 6.5 times as high as in the S strain. The increase in esterase activity was closely associated with the development of resistance in the R strains. DEF (S,S,S-tributyl phosphorotrithioate) proved to be an inhibitor for all esterases, with a particularly potent action on p-NPA-hydrolyzing enzymes. The inhibitory action was more pronounced in R strains than in the S strain. Pretreatment with DEF increased the toxicity of pyrethroid compounds in the R strains more than in the S strain and hence decreased the levels of resistance in these strains. This is evidence that the esterases contribute to the resistance against synthetic pyrethroids in S. littoralis larvae.  相似文献   

9.
The susceptibility to malathion, and the activity and sensitivity of acetylcholinesterase (AChE, EC 1.1.1.7) were compared between two populations of the oriental migratory locust, Locusta migratoria manilensis (Meyen) collected from Wudi County of Shandong Province in East China and Huangliu County of Hainan Province in South China. Huangliu population showed 8.5-fold resistance to malathion compared with Wudi population. AChE from Huangliu population showed 4.8-fold higher activity than that from Wudi population toward the model substrate acetylthiocholine (ATC). Kinetic studies indicated that AChE from Huangliu population had 2.6-fold lower affinity, but 5.0-fold higher catalytic activity toward ATC than AChE from Wudi population. Significantly increased activity of AChE in Huangliu population was also confirmed by non-denaturing polyacrylamide gel electrophoresis. Inhibition kinetics revealed that AChE from Huangliu population was 9.8-, 2.4-, 8.0- and 7.7-fold less sensitive to inhibition by paraoxon, malaoxon, chlopyrifos oxon, demeton-S-methyl, respectively, than that from Wudi population. Our studies revealed that a mild resistance to malathion in Huangliu population was associated with reduced sensitivity and increased catalytic activity of AChE. Our results suggest that alterations of AChE may play an important role conferring or contribute to malathion resistance in Huangliu population of the locust.  相似文献   

10.
Juvenile hormone III was tritium labeled on the methyl ester and utilized with other substrates in an investigation of inhibition and substrate specificity of hemolymph esterases from the cockroach, Blaberus giganteus. The structure of labeled juvenile hormone III was supported both chemically and biochemically. Forty-two potential inhibitors were examined, and the best inhibitors included phosphoramidothiolates and S-phenylphosphates. One of these inhibitors was found useful in hormone biosynthesis studies dealing with the enzymatic conversion of methyl farnesoate to juvenile hormone in corpora allata homogenates. Several commonly used inhibitors of carboxyesterases caused only weak inhibition of JH esterases. Gel filtration elution patterns, inhibitor relationships, and specific activities of the hemolymph esterases indicate that juvenile hormones I and III are degraded by similar if not identical enzymes. In some cases, α-naphthyl acetate and juvenile hormone esterase activity could be differentially inhibited. Hemolymph esterases were not capable of degrading ethyl or isopropyl conjugated esters of two juvenoids or three model substrates.  相似文献   

11.
Separate esterase activities of rat and mouse liver microsomes hydrolyzing malathion, trans-permethrin, and cis-permethrin were differentiated on the basis of their sensitivities to inhibition by paraoxon and α-naphthyl N-propylcarbamate (NPC). In rat liver microsomes, the malathionhydrolyzing activity was more sensitive to both inhibitors and showed a different time course of NPC inhibition than the activities hydrolyzing the permethrin isomers. Paraoxon completely inhibited trans-permethrin hydrolysis, but only partially inhibited that of cis-permethrin. The paraoxonsensitive trans- and cis-permethrin-hydrolyzing activities were not differentially inhibited, but separate inhibition curves were obtained for the inhibition of trans- and cis-permethrin hydrolysis by NPC. The mouse liver esterase activity hydrolyzing trans-permethrin showed a similar paraoxon sensitivity to that of rat liver, but that the paraoxon-sensitive portion of the cis-permethrinhydrolyzing activity was 5.5-fold less sensitive to paraoxon than the corresponding rat liver activity and was clearly differentiated from the mouse liver trans-permethrin-hydrolyzing activity. The mouse liver malathion-hydrolyzing activity was 100-fold less sensitive to paraoxon and 14-fold less sensitive to NPC than the corresponding rat liver activity. Rat and mouse liver esterase activities hydrolyzed trans- and cis-permethrin at similar rates under standard assay conditions, but mouse liver esterases were 10-fold less active in hydrolyzing malathion. The higher specific activity of rat liver malathion-hydrolyzing esterases resulted from the greater apparent affinity and maximum velocity for malathion hydrolysis. These results demonstrate that the hydrolysis of malathion, trans-permethrin, and cis-permethrin by rat and mouse liver microsomal preparations involves several esterases with differing substrate specificities and inhibitor sensitivities.  相似文献   

12.
通过生物测定表明高密棉蚜对有机磷的抗性高于北京棉蚜,用紫外分光光度计比色法(A法)及微量滴度酶标板法(B法 )测定高密棉蚜及北京棉蚜的α-乙酸萘酯酯酶活力和α-乙酸萘酯酯酶动力学。北京棉蚜和高密棉蚜的α- NA酯酶活力分别为 2.23、4.48(A法 )和1.13、3.30(B法)μmol·mg-1pro.·min-1,高密、北京棉蚜的酶活之比为 2 .00(A法 )、2 .92(B法) ;北京棉蚜、高密棉蚜的Km值分别为:6.06×10-5、7.51× 10-5(A法 )和 7.66×10-5、8.87×10-5 (B法) mol·L-1,Vmax值为2.53、5.82(A法)和1.28、3.61(B法)μmol·mg-1·min-1。比较紫外分光光度计比色法及微量滴度酶标板法的测定结果,表明微量滴度酶标板法的测定结果是可靠的。  相似文献   

13.
Malathion resistance of a field-collected population of Rhizopertha dominica (Coleoptera: Bostrichidae) from Mexico was evaluated and the resistance mechanisms were characterized both in vivo and in vitro. The Mexican population showed a resistance level of 50-fold at LC50 as compared with that of a susceptible laboratory population. Malathion bioassays with the synergists triphenyl phosphate, piperonyl butoxide and diethyl maleate suggested that esterases were likely to contribute to the resistance whereas cytochrome P450 monooxygenases and glutathione S-transferases were not. In-vitro assays of esterases indicated that the general esterase activity was 1·3-fold higher in the Mexican population than in the susceptible population. However, the phosphotriesterase activity in the resistant population was 3·7-fold higher than in the susceptible population. Significantly higher phosphotriesterase activity in the resistant population was further indicated by 3·4-fold increase of Vmax in enzyme kinetics and higher frequency of individuals with high phosphotriesterase activity in this population. All these findings suggested that phosphotriesterases play a role in malathion resistance in the Mexican population of lesser grain borer. © 1998 SCI  相似文献   

14.
Increased hydrolytic metabolism of organophosphate insecticides has been associated with resistance among Nebraska western corn rootworm populations. In this study, resistance-associated esterases were partially purified by differential centrifugation, ion exchange, and hydroxyapatite column chromatography, with a final purification factor of 100-fold and recovery of approximately 10%. Kinetic analysis of the partially purified enzyme indicated that the Km of the group II esterases was identical for the two populations, although Vmax was consistently threefold higher in the resistant population. A putative esterase, DvvII, was further purified to homogeneity by preparative polyacrylamide gel electrophoresis. DvvII is a monomer with a molecular weight of approximately 66 kDa, although three distinct isoforms with similar pIs were evident based on isoelectric focusing gel electrophoresis. Immunoassays with the Myzus persicae E4 antiserum indicated that group II esterases from D. v. virgifera were cross-reactive and expressed at much higher titers in the resistant population relative to the susceptible counterpart. These results suggest that the resistance is likely associated with overproduction of an esterase isozyme in resistant D. v. virgifera populations.  相似文献   

15.
The role of esterase in pyrethroid resistance was studied in the final larval instar of different strains of the cotton bollworm, Helicoverpa armigera. The resistant strains viz., Nagpur strain and the Delhi strain were found to have elevated midgut esterase activity in comparison to the susceptible strain. Nagpur strain and Delhi strain have 2.24 and 1.73-fold higher esterase activity, respectively, than that of the susceptible strain. The Native PAGE displayed important differences in the midgut esterase isozyme pattern between the susceptible and the pyrethroid-resistant strains. Out of the 10 esterase isozyme observed, susceptible strain lacked three bands, E2, E6 and E10 that were found in the resistant strains. The potency of the synergists piperonyl butoxide (PBO) and dihydrodillapiole (DDA) as esterase inhibitor were also studied both in vitro and in vivo. The in vitro results clearly show that both PBO and DDA inhibited esterase activity in the two resistant strains, while there was almost no esterase inhibition in the homogenate of the susceptible strain. The in vivo inhibition studies (topical application of PBO and DDA followed by biochemical analysis) illustrated that PBO- and DDA-esterase binding is rather slow and non permanent process. Esterase inhibition did not occur immediately after the synergist treatment but at 4 and 8 h post treatment in case of PBO and DDA, respectively. Native PAGE revealed that the in vivo esterase inhibition caused by both PBO and DDA was due to the binding of the synergist with the E6 isozyme which was not present in the susceptible strain.  相似文献   

16.
Despite the frequent and widespread applications of organophosphates against Cydia pomonella this species has developed low levels of resistance to this chemical group. Investigations concerning the mechanisms involved in resistance are scarce, and usually consider only one of the potential mechanisms. With the aim of a better understanding the resistance mechanisms and their possible interaction, four of these mechanisms were investigated simultaneously in one sensitive (Sv) and two resistant strains (Raz and Rdfb) of this insect. Resistant strains displayed an increased mixed function oxidase activity, whereas carboxylesterase activity varied upon the substrate used. The three strains had similar β-naphtyl acetate activity, and the hydrolysis of α-naphthyl acetate and p-nitrophenyl valerate was higher in the Sv strain. The p-nitrophenyl acetate activity was highest in the resistant strains and was strongly inhibited by azinphos and DEF. The Raz strain has a modified acetylcholinesterase (AChE), which resulted in a 0.7-, 3.2- and 21.2-fold decrease in the susceptibility to chlorpyriphos-ethyl-oxon, azinphos-methyl-oxon, and paraoxon-methyl, respectively. These combined resistance mechanisms only conferred to Raz a 0.6-, 7.9- and 3.1-fold resistance to the related insecticides. Organophosphates resistance in C. pomonella results from a combination of mechanisms including modified affinities to carboxylesterase substrates, and increased metabolisation of the insecticide. The apparent antagonism between increased functionalisation and reduced sensitivity of the AChE target is discussed.  相似文献   

17.
Malathion resistance in Anopheles stephensi from Pakistan was synergized by triphenyl phosphate, primarily a carboxylesterase inhibitor. There was a slight degree of antagonism with piperonyl butoxide. The major metabolite of malathion in larvae of both the resistant and susceptible strains was malathion monocarboxylic acid. Resistant larvae produced about twice as much of this product as the susceptible larvae. This suggests that a qualitative or a quantitative change in a carboxylesterase enzyme may be the basis of malathion resistance in this strain. Analysis of general esterase levels to α- and β-naphthyl acetate showed that there was no quantitative change in the amount of carboxylesterase enzyme present in the resistant strain as compared to the susceptible.  相似文献   

18.
BACKGROUND: Frankliniella occidentalis (Pergande) is among the most important crop pests in the south‐east region of Spain; its increasing resistance to insecticides constitutes a serious problem, and understanding the mechanisms involved is therefore of great interest. To this end, F. occidentalis populations, collected from the field at different locations in south‐east Spain, were studied in terms of total esterase activity and esterase isoenzyme pattern. RESULTS: Individual thrips extracts were analysed by native polyacrylamide gel electrophoresis (PAGE) and stained for esterase activity with the model substrate α‐naphthyl acetate. Significant correlations were found between resistance to the insecticides acrinathrin and methiocarb and the presence of a group of three intensely stained bands, named Triplet A. For each individual thrips extract, total esterase activity towards the substrates α‐naphthyl acetate and α‐naphthyl butyrate was also measured in a microplate reader. Insects possessing Triplet A showed a significantly higher α‐naphthyl acetate specific activity and α‐naphthyl acetate/α‐naphthyl butyrate activity ratio. This observation allowed a reliable classification of susceptible or resistant insects either by PAGE analysis or by total esterase activity determination. CONCLUSION: The PAGE and microplate assays described can be used as a monitoring technique for detecting acrinathrin‐ and methiocarb‐resistant individuals among F. occidentalis field populations. Copyright © 2008 Society of Chemical Industry  相似文献   

19.
The hydrolysis of trans-permethrin and α-naphthyl acatate by midgut, fat body, and cuticle homogenates from Pseudoplusia includens (Walker) was monitored during the development of the last instar. The midgut homogenates appeared to have two pH optima (7.6 and 8.6) for the hydrolysis of trans-permethrin, the fat body homogenates had one optimum (7.4–7.8), and the cuticle homogenates had a major optimum at 6.6. Hydrolysis of both substrates peaked during the late feeding stages for midgut and cuticle homogenates, although relative changes were not the same. Hydrolysis of trans-permethrin peaked during the late feeding stage in fat body homogenates, while hydrolysis of α-naphthyl acetate continually increased through the prepupal stage. Thus, the hydrolysis of α-naphthyl acetate is not necessarily associated with the hydrolysis of trans-permethrin. The LD50 values for trans-permethrin on the different stages appeared to reflect the influence of hydrolysis.  相似文献   

20.
The role of esterases as related to insecticide resistance was studied in an organophosphorus (OP)-resistant strain of the green rice leafhopper. As judged by p-nitrophenyl acetate hydrolysis, 21, 5, and 74% of the esterase activity was located in nuclei/mitochondria, microsomes, and the soluble fraction, respectively. All the fractions were active in hydrolyzing malathion, paraoxon, and fenvalerate. Hydrolysis of malathion and fenvalerate increased with time while that of paraoxon reached a plateau within 15 min. Since a considerable amount of p-nitrophenol was detected in the paraoxon reaction at 0°C and at zero time, the formation of p-nitrophenol may be due to phosphorylation of the esterases rather than phosphorotriesterase action. The results suggest a dual role for esterases in resistance mechanisms; a catalyst for hydrolysis of malathion and fenvalerate, and a binding protein for the oxygen analogs of other OP insecticides, both of which would protect the intrinsic target, acetylcholinesterase, from inhibition. Chromatofocusing of the soluble fraction resolved five esterase peaks, I–V. These esterases were active toward the three general substrates as well as for the three insecticides tested, except for Peak I in which the overall activity was too low. Thin-layer agar gel electrophoresis showed that the chromatofocusing peaks I–V corresponded to the electrophoretic bands E1–E5, some of which were previously shown to be associated with OP resistance. The dual role of these esterases may explain the cross-resistance between malathion and other OP insecticides as well as synergism between OP and carbamate insecticides.  相似文献   

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