A biochemical mechanism of resistance to pirimicarb in two glasshouse clones of Aphis gossypii

The susceptibility of three clones of Aphis gossypii Glover to 15 insecticides was established by bioassay. A high level of resistance towards pirimicarb was confirmed for a clone from Holland (Dutch R) and a clone from Japan (Jap R), while the susceptible clone (S) was killed by very low doses of the insecticide. However, only limited cross-resistance was shown towards other carbamates and organophosphates, and no marked resistance to the pyrethroids tested. The acetylcholinesterase (AChE) of both resistant clones hydrolysed acetyl-choline faster than that of susceptible aphids, with greatest enzyme activity shown by the Dutch R clone. Inter-clone differences in these rates were consistent with differences in catalytic centre activities. Inhibition (I₅₀) of AChE by pirimicarb was approximately 900-fold higher for the resistant clones than for the S clone. First-order kinetics revealed that resistance to pirimicarb in Dutch R and Jap R involved a modified AChE which had a reduced (approximately 350-fold) affinity (K_d) for pirimicarb. The marked change in AChE affinity for pirimicarb was not repeated with the other carbamates tested, ethiofencarb and aldicarb. It was considered that the resistant aphids would not require mechanisms in addition to insensitive AChE in order to show the high level of resistance to pirimicarb shown in the bioassay.     

Resistance mechanisms and associated mutations in acetylcholinesterase genes in Sitobion avenae (Fabricius)

Wheat aphid, Sitobion avenae (fabricius), is one of the most important wheat pests and has been reported to be resistant to commonly used insecticides in China. To determine the resistance mechanism, the resistant and susceptible strains were developed in laboratory and comparably studied. A bioassay revealed that the resistant strain showed high resistance to pirimicarb (RR: 161.8), moderate reistance to omethoate (32.5) and monocrotophos (33.5), and low resistance to deltamethrin (6.3) and thiodicarb (5.5). A biochemistry analysis showed that both strains had similar glutathione-S-transferase (GST) activity, but the resistant strain had 3.8-fold higher esterase activity, and its AChE was insensitive to this treatment. The I₅₀ increased by 25.8-, 10.7-, and 10.4-folds for pirimicarb, omethoate, and monocrotophos, respectively, demonstrating that GST had not been involved in the resistance of S. avenae. The enhanced esterase contributed to low level resistance to all the insecticides tested, whereas higher resistance to pirimicarb, omethoate, and monocrotophos mainly depended on AChE insensitivity. However, the AChE of the resistant strain was still sensitive to thiodicarb (1.7-fold). Thus, thiodicarb could be used as substitute for control of the resistant S. avenae in this case. Furthermore, the two different AChE genes cloned from different resistant and susceptible individuals were also compared. Two mutations, L436(336)S in Sa.Ace1 and W516(435)R in Sa.Ace2, were found consistently associated with the insensitivity of AChE. They were thought to be the possible resistance mutations, but further work is needed to confirm this hypothesis.     

Insecticide resistance in cotton aphid Aphis gossypii (Glov.) in the Sudan Gezira

The resistance to insecticides of three Sudanese strains of A. gossypii (Glov.) collected from cotton fields in the Sudan Gezira Scheme over three seasons (1988, 1989, 1990) and that of two French strains was studied in the laboratory. When compared with a known susceptible strain, the aphids were found to be resistant-to the eight insecticides tested. Evolution of resistance in Sudanese strains during the three crop seasons was observed. Assay of aphid homogcnate for carboxytesterase activity towards the substrates α-naphthyl acetate and β-naphthyl acetate showed that there was no enhancement of this class of enzyme and thus it was not a cause of resistance in this species. A study of interaction between the acetylcholinesterase (AchE) and pirimicarb established the kinetics of the inhibition process. I₅₀ values were found to be much higher for the Sudanese strains than for the susceptible strain. First-order inhibition kinetics revealed that resistance towards pirimicarb in Sudanese-strains was caused by a modified AchE which had a reduced affinity (higher K_a value) and poor carbamylation ability (lower K₂ value) for pirimicarb. The resistance mechanisms for the other insecticides remain to be studied.     

药剂对小菜蛾抗性及敏感品系乙酰胆碱酯酶抑制作用比较

采用浸叶法测定了云南通海、元谋和澜沧的小菜蛾plutella xylostella田间种群对常用杀虫剂的抗药性。结果表明,云南上述地区小菜蛾田间种群对各类杀虫剂均产生了不同程度的抗性。对有机磷类药剂的抗药性为1.74~31.1倍;对菊酯类药剂的抗药性为7.41~764倍;对阿维菌素类药剂则产生了 5.60~4.06×104倍的抗性。通过离体和活体试验测定了药剂对小菜蛾头部乙酰胆碱酯酶(AChE)的抑制作用。敌敌畏和灭多威对通海抗性品系AChE离体和活体内的抑制中浓度(I50)分别是敏感品系的209、26.5倍和2.21、2.16倍;敌敌畏对通海小菜蛾种群的离体和活体内抑制中时间(IT50)小于敏感品系,分别是敏感品系的0.32和0.17倍;而灭多威对通海小菜蛾种群的离体和活体内抑制中时间(IT50)则大于敏感品系,分别是敏感品系的1.37和1.74倍。     

Note: Insecticide resistance in<Emphasis Type="Italic">Helicoverpa armigera</Emphasis> from cotton-growing areas in Turkey

Insecticide resistance inHelicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) is one of the most important constraints on cotton production in Turkey. We investigated the susceptibility ofH. armigera strains collected from cotton fields in the Adana, Hatay and Antalya provinces to insecticides which are in wide use. LD₅₀ values for tralomethrin, lambda-cyhalothrin, endosulfan, profenofos and methomyl were determined by topical bioassay. At the LD₅₀ levels, resistance ratios for tralomethrin were 24.7-, 19.7- and 15.7-fold in the Adana, Hatay, and Antalya strains, respectively; and for lambda-cyhalothrin were 41-, 20-and 40-fold, respectively. Resistance ratios ranged from 1.2- to 2.1-fold in all field strains for endosulfan, profenofos and methomyl, with no significant resistance. These results suggest the presence of resistance to tralomethrin and lambda-cyhalothrin inH. armigera strains collected from cotton fields, but no resistance to endosulfan, profenofos or methomyl could be observed.     

江苏省水稻二化螟的抗药性测定及对甲胺磷的抗性机制

采用3龄幼虫点滴法测定了江苏省水稻二化螟对杀虫单和甲胺磷的抗性水平，并就二化螟对甲胺磷产生抗性的机制进行了研究，结果表明，高淳和徐州二化螟种群对杀虫单已达高抗水平，其它地区也表现为中等抗生，在所测定的6个种群中，苏州种群对甲胺磷处于低抗水平，其它种群为中抗种群，澌知体增效作用表明，氧化胡椒基丁醚（PBO）对甲胺磷有明显的增效作用，磷酸三苯酯（TPP）和顺丁烯酸二乙酸（DEM）的增效作用不明显，对离体解毒酶的研究表明，R种群幼虫多功能氧化酶（MFO）的O-脱甲基作用的活力比S幼虫高，R幼虫乙酰胆碱酯酶（AChE)的活力与S幼虫无显著差异。以甲胺磷为抑制剂的R幼虫I50是S幼虫的2.52倍，二化螟对甲胺磷的抗性机理可能与MFO的O-脱甲基作用活力增矿山和AChE的敏感性降低有关。     

Biochemical evidence that an S431F mutation in acetylcholinesterase-1 of Aphis gossypii mediates resistance to pirimicarb and omethoate

Molecular changes in acetylcholinesterase (AChE) leading to target-site resistance to carbamate and organophosphate insecticides have recently been identified. Of particular interest is the S431F mutation in ace2 and its orthologue ace1 of Myzus persicae Sulzer and Aphis gossypii Glover, respectively. This mutation has been correlated with resistance to pirimicarb, but biochemical evidence has not yet been provided. Here, we describe for the first time that recombinantly expressed AChE1 from A gossypii carrying the S431F mutation is insensitive to pirimicarb and omethoate, but sensitive to demeton-S-methyl and hypersensitive to carbofuran. Furthermore, site-directed mutagenesis of the serine residue at position 431 in ace1 from a pirimicarb-susceptible clone of A gossypii conferred insensitivity to pirimicarb. We conclude that AChE1 of A gossypii is the target of toxicological relevance of carbamates and organophosphates.     

不同体色瓜蚜耐药性差异及其生化机制

试验结果表明,在黄瓜上的绿色瓜蚜(Aphis gossypii Glov.)种群对氧化乐果、抗蚜威和氰戊菊酯的耐性明显高于黄色瓜蚜种群。这两种颜色的瓜蚜乙酰胆碱酯酶(ACHE)具有明显的不同,在绿色种群中,有60.7%的个体AChE活性大于0.8(OD_(412)/mg蛋白质,分钟),而在黄色种群中则仅有25.3%的个体AChE活性大于0.8;氧化乐果(终浓度为1.69×10~(-2)mol/L)对单个蚜虫AChE的抑制率,前者仅有19%的个体AChE抑制率在30%以上,而后者则有46%的个体在30%以上。α-乙酸萘酯羧酸酯酶的活性大于25(×10~(-2)μmol/蚜,15分钟)的个体在绿色种群中占56%,而在黄色种群中仅有20%;β-乙酸萘酯羧酸酯酶活性大于25(×10~(-2)μmol/蚜,15分钟)的个体在绿色瓜蚜中占45%,而在黄色种群中仅有7%。说明绿色和黄色瓜蚜耐药性的差异可能是由于AChE敏感性和羧酸酯酶活性的不同造成的。     

Acetylcholinesterase point mutations in European strains of Tetranychus urticae (Acari: Tetranychidae) resistant to organophosphates

BACKROUND: In Tetranychus urticae Koch, acetylcholinesterase insensitivity is often involved in organophosphate (OP) and carbamate (CARB) resistance. By combining toxicological, biochemical and molecular data from three reference laboratory and three OP selected strains (OP strains), the AChE1 mutations associated with resistance in T. urticae were characterised. RESULTS: The resistance ratios of the OP strains varied from 9 to 43 for pirimiphos‐methyl, from 78 to 586 for chlorpyrifos, from 8 to 333 for methomyl and from 137 to 4164 for dimethoate. The insecticide concentration needed to inhibit 50% of the AChE1 activity was, in the OP strains, at least 2.7, 55, 58 and 31 times higher for the OP pirimiphos‐methyl, chlorpyrifos oxon, paraoxon and omethoate respectively, and 87 times higher for the CARB carbaryl. By comparing the AChE1 sequence, four amino acid substitutions were detected in the OP strains: (1) F331W (Torpedo numbering) in all the three OP strains; (2) T280A found in the three OP strains but not in all clones; (3) G328A, found in two OP strains; (4) A201S found in only one OP strain. CONCLUSIONS: Four AChE1 mutations were found in resistant strains of T. urticae, and three of them, F331W, G328A and A201S, are possibly involved in resistance to OP and CARB insecticides. Among them, F331W is probably the most important and the most common in T. urticae. It can be easily detected by the diagnostic PCR‐RLFP assay developed in this study. Copyright © 2009 Society of Chemical Industry     

GABA receptors of insects susceptible and resistant to cyclodiene insecticides

Toxicity tests revealed up to 40-fold resistance to a number of cyclodiene insecticides in a laboratory-reared, cyclodiene-resistant (CYW) housefly strain (Musca domestica L.). Using [³⁵S] TBPS as a probe for convulsant sites in insects, saturable specific binding was detected in thorax and abdomen membranes prepared from housefly strains susceptible (CSMA) and resistant (CYW) to cyclodienes. Scatchard analysis of[³⁵S] TBPS binding data to CSMA and CYW membranes failed to provide evidence for significant differences between the two strains in either the affinity (K_d) or density (B_max) of saturable binding sites. For several polychlorocycloalkane insecticides, the ligand displacement profile of [³⁵S] TBPS binding was almost identical for the CSMA and CYW houseflies. Therefore, using [³⁵S] TBPS as a probe for convulsant sites, a 40-fold resistance to cyclodienes in the CYW housefly strain cannot be accounted for only in terms of alterations in TBPS binding sites.     

Resistance risk assessment of two insect development inhibitors,diflubenzuron and cyromazine,for control of the housefly Musca domestica. part i: Larvicidal tests with insecticide-resistant laboratory and danish field populations

Larvicide tests with diflubenzuron (DFB) and cyromazine (CYR) were carried out against 15 laboratory housefly strains and 89 field strains collected from 87 Danish and 2 Swedish farms, 1975–89. The strains represented a wide range of adult insecticide resistance and R-mechanisms. The larvicide tests were done by treating larval medium with serial or discriminating dosages of the larvicide, seeding it with eggs and calculating the mortality during development to adults. The WHO susceptible strain (S) was used as a reference. Dose-response tests with DFB gave resistance ratios (R/S) from 1·1 to 4·1 at LC₅₀ and 0·3 to 3·4 at LC₉₅ and, with CYR, R/S from 0·6 to 1·8 at LC₅₀ and 0·6 to 2·9 at LC₉₅. It was concluded that the relatively small variation in susceptibility between strains was not generally correlated with resistance in adult flies to organophosphorus, pyrethroid or other conventional insecticides (neurotoxins). Tests with discriminating dosages of DFB (59 farms) and CYR (63 farms) showed no indication of resistance to either product. The results of investigations by other workers on the relation between resistance to DFB or CYR and resistance to conventional insecticides are discussed.     

Triazophos resistance mechanisms in the rice stem borer (Chilo suppressalis Walker)

A field population of the rice stem borer (Chilo suppressalis Walker) with 203.3-fold resistance to triazophos was collected. After 8-generation of continuous selection with triazophos in laboratory, resistance increased to 787.2-fold, and at the same time, the resistance to isocarbophos and methamidophos was also enhanced by 1.9- and 1.4-fold, respectively, implying some cross-resistance between triazophos and these two organophosphate insecticides. Resistance to abamectin was slightly enhanced by triazophos selection, and fipronil and methomyl decreased. Synergism experiments in vivo with TPP, PBO, and DEM were performed to gain a potential indication of roles of detoxicating enzymes in triazophos resistance. The synergism results revealed that TPP (SR, 1.92) and PBO (SR 1.63) had significant synergistic effects on triazophos in resistant rice borers. While DEM (SR 0.83) showed no effects. Assays of enzyme activity in vitro demonstrated that the resistant strain had higher activity of esterase and microsomal O-demethylase than the susceptible strain (1.20- and 1.30-fold, respectively). For glutathione S-transferase activity, no difference was found between the resistant and the susceptible strain when DCNB was used as substrate. However, 1.28-fold higher activity was observed in the resistant strain when CDNB was used. These results showed that esterase and microsomal-O-demethylase play some roles in the resistance. Some iso-enzyme of glutathione S-transferase may involve in the resistance to other insecticides, for this resistant strain was selected from a field population with multiple resistance background. Acetylcholinesterase as the triazophos target was also compared. The results revealed significant differences between the resistant and susceptible strain. The V_max and K_m of the enzyme in resistant strain was only 32 and 65% that in the susceptible strain, respectively. Inhibition tests in vitro showed that I₅₀ of triazophos on AChE of the resistant strain was 2.52-fold higher. Therefore, insensitive AChE may also involved in triazophos resistance mechanism of rice stem borer.     

Note: Pyrethroid resistance and possible involvement of glutathione<Emphasis Type="Italic">S</Emphasis>-transferases in<Emphasis Type="Italic">Helicoverpa armigera</Emphasis> from Turkey

An introductory study was conducted to investigate the pyrethroid resistance ofHelicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) strains in Turkey, collected from cotton fields in the Adana and Antalya provinces, through two different synthetic pyrethroid insecticides: lambda-cyhalothrin and esfenvalerate. In addition, the roles of glutathioneS-transferases (GSTs) in this resistance mechanism were analyzed. It was found that whereas resistance ratios for lambda-cyhalothrin (LD₅₀ levels) were 3- and 98-fold increased in the Adana and Antalya strains, respectively, esfenvalerate ratios were 3.3- and 92.3-fold increased in the Adana and Antalya strains, respectively, with respect to the susceptible strain. Furthermore, Adana and Antalya strains showed 2.4- and 2.9-fold higher GST activities than the susceptible strain, respectively. In the Antalya field strain, the minor increase in GST activity compared with the resistance levels implies that GSTs may be not greatly involved in this resistance. It also provides evidence that they could not be the only metabolic mechanism responsible for resistance to lambda-cyhalothrin and esfenvalerate inH. armigera from Turkey. http://www.phytoparasitica.org posting Nov. 16, 2006.     

Toxicological and biochemical characterizations of AChE in Liposcelis bostrychophila Badonnel (Psocoptera: Liposcelididae)

The toxicological and biochemical characteristics of acetylcholinesterases (AChE) in the resistant and susceptible strains (SS) of Liposcelis bostrychophila were investigated. The two resistant strains were the dichlorvos-resistant strain (DDVP-R) and the phosphine-resistant strain (PH₃-R) with resistance ratios of 22.36 and 4.51, respectively. Compared to their susceptible counterpart, the AChE activity per insect and the specific activity of AChE in DDVP-R and PH₃-R were significantly higher. There were also significant kinetic differences between DDVP-R and PH₃-R. The apparent Michaelis-Menten constant (K_m) for acetylthiocholine iodide (ATChI) was obviously lower in SS than that in PH₃-R, indicating a higher affinity to the substrate ATChI in the susceptible strains. The affinity for the substrate ATChI in DDVP-R and SS were not significantly different. The V_max value of the PH₃-R was significantly greater when compared to the V_max for the SS suggesting a possible over expression of AChE in this resistant strain. The inhibition of AChE to insecticide exposure in vitro revealed that all six insecticides were inhibitory for the extracted AChE’s. Based on the I₅₀ values, AChE of the SS were more sensitive to dichlorvos, paraoxon-ethyl, malaoxon and demeton-S-methyl than those of the two resistant strains. As for carbaryl and eserine, the PH₃-R suggested a significantly higher I₅₀s compared to the susceptible strain, while, no significant differences were found between SS and DDVP-R.     

Field-simulator study of insecticide resistance conferred by esterase-, MACE- and kdr-based mechanisms in the peach-potato aphid,Myzus persicae (Sulzer)

Insecticide sprays were applied to Myzus persicae (Sulzer) populations carrying various combinations of three insecticide resistance mechanisms (esterase-based metabolic resistance and two target site mechanisms, known as MACE and kdr), supported on host plants growing in field simulator cages. The study showed that MACE confers extreme resistance to pirimicarb and triazamate (carbamate insecticides) but not to deltamethrin + heptenophos (16 + 1) (Decisquick) or dimethoate (an organophosphorus insecticide). Resistance to dimethoate depends solely on levels of esterase-based resistance, while resistance to Decisquick depends on kdr and esterase. None of the four insecticides is effective against aphids carrying MACE combined with extreme esterase-based resistance. This knowledge, in association with current monitoring of the mechanisms, will play an important role in making decisions on insecticide use against M persicae in the UK. © 1999 Society of Chemical Industry     

Insecticide resistance status of the codling moth Cydia pomonella (Lepidoptera: Tortricidae) from Greece

The codling moth Cydia pomonella L. is controlled mostly with chemical insecticides in Greece and control failures have been reported. However, there are no insecticide resistance studies in the country as yet. We examined the insecticide resistance status of 33 and 38 populations of fifth-instar non-diapausing and diapausing larvae, respectively by applying bioassays, biochemical and DNA diagnostics. Diagnostic concentrations of azinphos-methyl, phosalone, deltamethrin, thiacloprid, fenoxycarb, tebufenozide, methoxyfenozide and diflubenzuron were used in bioassays. Almost all populations showed reduced susceptibility to at least one insecticide and approximately half of them to all insecticides examined compared to a laboratory susceptible strain used as reference. However, only one out of six populations tested showed reduced susceptibility in ovicidal tests with fenoxycarb. Cross-resistances were observed among most insecticides, except from the pairs fenoxycarb–phosalone and thiacloprid–phosalone, in non-diapausing larvae. The more obvious biochemical marker associated with the reduced susceptibility observed in both larval instars was elevated cytochrome P₄₅₀ polysubstrate monooxygenases activity, followed by elevated glutathione-S-transferase activity and reduced carboxylesterases activity. Neither sodium channel nor AChE known resistance mutations were found in any of the approximately 1000 individuals of each larval instar screened with diagnostic PCR. Actions for Integrated Resistance Management and application of alternative control methods are discussed.     

The pattern of cross-resistance to insecticides and juvenile hormone analogues in a diflubenzuron-resistant strain of the cotton leaf worm Spodoptera littoralis boisd

The rate of development of resistance to diflubenzuron in a laboratory susceptible strain of the cotton leafworm Spodoptera littoralis Boisd., the pattern of cross-resistance exhibited by the resistant strain to several insecticides and juvenile hormone analogues, as well as the synergistic action of piperonyl butoxide (PB) and S, S, S-tributyl phosphorotrithioate (TBP) with insecticides or diflubenzuron on both strains, were investigated. Resistance to diflubenzuron increased slightly in the first eight selected generations and was enhanced by further selection until in generation 30, the selected strain attained the high level of resistance of 290.7-fold, compared with the parent strain. The resistant strain when challenged with either insecticides or juvenile hormone analogues at selected generations (5, 10, 15, 20, 25 and 30) exhibited different levels of resistance to several insecticides representing organochlorine and organo-phosphorus compounds, carbamates and pyrethroids, but a clear case of negative correlation was indicated between resistance to diflubenzuron and juvenile hormone analogues. With regard to the synergistic action of PB and TBP on the toxicity of either diflubenzuron or insecticides against the fourth-instar larvae of the susceptible strain, methomyl showed slight levels of synergism when it was combined with them. With the exception of cypermethrin, which was not affected by the two synergists, lower levels of synergism were observed with the compounds endrin, diflubenzuron and fenvalerate when they were combined with the same synergists. These two synergists however, antagonised the toxic action of the organophosphorus compounds phosfolan and chlorpyrifos. Against the resistant strain, endrin was moderately synergised by TBP but only slightly by PB. Slight levels of synergism were observed when methomyl, phosfolan and diflubenzuron were combined with either synergist, but both antagonised chlorpyrifos and fenvalerate. Resistance to diflubenzuron and to the other tested chemicals in the resistant strain was scarcely affected by the two synergists.     

The detection and distribution of organophosphorus and carbamate insecticide-resistant Myzus persicae (Sulz.) in Britain in 1976

Examination of the enzyme that determines the level of resistance to organophosphorus insecticides and carbamates in Myzus persicae (Sulz.) and bioassays were used to establish the frequency and resistance levels of resistant aphids on outdoor crops in Britain in 1976. The biochemical tests, staining esterase-4 after electrophoresis and total esterase determination, were more sensitive than bioassays. However the dip-test, a simple, rapid and inexpensive bioassay designed to detect resistance and its different levels gave satisfactory results which warrant its use where biochemical detection of resistance is not possible. Carboxylesterase activities of M. persicae collected in 1976 fell into three groups: low, moderate and high, and these were correlated with differences in tolerance to dimethoate, demeton-S-methyl and pirimicarb. Aphids with low esterase activity were susceptible(S). Those with the moderately active enzyme (R₁) had five- to seven-fold resistance to the two organophosphorus insecticides and were marginally resistant (about two-fold) to pirimicarb. The insects with the most active esterase (R₂) were strongly resistant to dimethoate (resistance factor, RF × 126) and moderately resistant to demeton-S-methyl (RF × 17) and pirimicarb (RF × 8). Some R₁, but no S aphids survived the recommended dose of demeton-S-methyl on field crops probably because they were under the lowest leaves and therefore protected from direct contact with the spray. Laboratory tests demonstrated that these R₁ aphids tolerated the residual deposit and systemic dose present in the leaves of the treated potato-plants. This enabled their numbers to recover in treated fields much faster than the susceptible insects which could do so by immigration only when the residual dose in the plant was no longer toxic. R₁ aphids were common throughout the country particularly in eastern England where susceptible aphids were rare, but in the Shardlow area of Derbyshire susceptible aphids were in the majority throughout the summer. R₂ aphids were found only in samples from the west of Scotland and northern England. The implications of the presence of aphids with different levels of resistance for aphid control are discussed.     

The status and development of insecticide resistance in Danish populations of the housefly Musca domestica L

Samples of housefly (Musca domestica) field populations were collected from Danish livestock farms in 1997. The tolerance of the first‐generation offspring was determined for a number of insecticides. Dose‐response values were obtained by topical application for the pyrethroids bioresmethrin and pyrethrum, both synergised with piperonyl butoxide, and the organophosphate dimethoate. The organophosphates azamethiphos and propetamphos and the carbamate methomyl were tested in discriminating dose feeding bioassays. Resistance was low to moderate in most of the populations for most of the compounds tested, but this study also revealed the existence of high resistance to pyrethroid, organophosphate and carbamate insecticides in some populations. The resistance factors at LD₅₀ for bioresmethrin/piperonyl butoxide ranged between 2 and 98, and for pyrethrum/piperonyl butoxide between 2 and 29. Our results indicate that pyrethroid resistance in Denmark is increasing, since four of the 21 farms showed more than 100‐fold resistance at LD₉₅, a level of resistance only observed once before. Resistance factors at LD₅₀ for dimethoate ranged from 9 to 100, and showed two distinct trends: populations with either decreasing or increasing resistance. Resistance to azamethiphos was found to be widespread and high. Although two strains with high methomyl and propetamphos resistance were observed, methomyl and propetamphos resistance is moderate and appears not to be increasing. © 2001 Society of Chemical Industry