Cross-resistance between acylurea insect growth regulators in a strain of Plutella xylostella L. (Lepidoptera: Yponomeutidae) from Malaysia

The activities of the acylurea insect growth regulators, chlorfluazuron, teflubenzuron and difubenzuron, and the neurotoxic macrocyclic lactone, abamectin were assessed against a laboratory susceptible (FS) strain and a field (Cameron Highlands, Malaysia (CH)) strain of the diamondback moth, Plutella xylostella L. using a leaf-dip bioassay at 20°C. The time taken to achieve end-point mortality was found to vary considerably (9–17 days), being fastest with abamectin against the FS strain and slowest with difubenzuron against the CH strain. The order of activity (LC₅₀ at F6/7) against second-instar larvae of both strains was: abamectin > chlorfluazuron = teflubenzuron ? difubenzuron. Subsequent assays (F14) with the acylureas, flufenoxuron and hexaflumuron against the FS strain suggested that the former was slightly more active than chlorfluazuron or teflubenzuron, the latter slightly less active. The CH population was found to be 12.6-, 6.7-, 6.4- and 2.3-fold less sensitive to difubenzuron, teflubenzuron, chlorfluazuron and abamectin respectively than the FS strain. Selection of sub-populations of the CH strain with chlorfluazuron (CHL-SEL) and teflubenzuron (TEF-SEL) for six generations (F6-11), resulted in LC₅₀ resistance ratios of 109- and 315-fold respectively when compared with the FS strain, equivalent to an 18- and a 46-fold increase in resistance compared with the unselected CH strain. Marked cross-resistance was also demonstrated between chlorfluazuron and teflubenzuron in both sub-populations. However, there was no evidence of cross-resistance to dijlubenzuron and abamectin and little or no cross-resistance to flufenoxuron and hexaflumuron. Resistance to chlorfluazuron and teflubenzuron appeared to be relatively unstable in the TEF-SEL compared with the CHL-SEL sub-population (over 6–9 generations). However, reselection of the TEF-SEL population with chlorfluazuron (F18–20) led to a very rapid increase in resistance to chlorfluazuron and particularly teflubenzuron. For the latter compound, resistance factors of about 1000000 were obtained (F19, 21). Such values are probably only semi-quantitative, as above a certain level of resistance feeding bioassays with acylureas (compounds which are active to a significant extent by ingestion) are likely to become rate-limiting.     

Aqueous solubilities,photolysis rates and partition coefficients of benzoylphenylurea insecticides

The aqueous solubilities and octanol–water partition coefficients (K_OW) of the benzoylphenylurea (BPU) insecticides teflubenzuron, chlorfluazuron, flufenoxuron and hexaflumuron were determined in comparison with the more extensively studied diflubenzuron. Both teflubenzuron and hexaflumuron were substantially less water‐soluble (9.4 (± 0.3) µg litre⁻¹ and 16.2 (± 0.5) µg litre⁻¹ in water, respectively) than the value previously reported for diflubenzuron (89 (± 4) µg litre⁻¹ in water). Log K_OW values for diflubenzuron, teflubenzuron, hexaflumuron, flufenoxuron and chlorfluazuron were 3.8, 5.4, 5.4, 6.2 and 6.6, respectively, as determined using reverse‐phase HPLC. Photodegradation of hexaflumuron, teflubenzuron and diflubenzuron in water indicated hexaflumuron to be the most rapidly degraded of the three compounds at pH 7.0 (t_1/2 = 8.6 (± 0.4) h) and pH 9.0 (t_1/2 = 5 (± 1) h); diflubenzuron was the slowest of the three pesticides to degrade in pH 7.0 (17 (± 4) h) and pH 9.0 (8 (± 2) h) buffered water. In a solar simulator using river water buffered to pH 9.0, teflubenzuron, hexaflumuron and diflubenzuron half‐lives were 20 (± 4), 15 (± 2) and 12 (± 1) h, respectively; dark controls showed no loss of parent BPU over similar time periods. © 2000 Society of Chemical Industry     

Persistence of insect growth regulators against the rice weevil,sitophilus oryzae,in grain commodities

Ten insect growth regulators (IGRs): four juvenile hormone analogs (JHAs) — fenoxycarb, MV-678, R-20458 and dofenapyn, and six molt inhibitors (MIs) — triflumuron, chlorfluazuron, hexaflumuron, teflubenzuron, Dowco 439 and flufenoxuron; and the organophosphate malathion, were evaluated for residual activity in stored grain for 2 years against the rice weevil,Sitophilus oryzae (L.). Triflumuron, flufenoxuron, teflubenzuron, chlorfluazuron, hexaflumuron and fenoxycarb, each at a concentration as low as 1 ppm a.i., were superior to 10 ppm malathion throughout the experiment. The six MIs, each tested at 50 and 100 ppm a.i., gave complete control for 2 years. Moreover, a concentration as low as 1 ppm a.i. of triflumuron, teflubenzuron, flufenoxuron and chlorfluazuron had good residual activity for 1 year post-treatment and provided almost complete grain protection during this period. Among the JHAs, fenoxycarb had the best residual activity. It gave almost complete control (96.6%) ofS. oryzae at 10 ppm for 18 months post-treatment. The other JHAs at the four test concentrations failed to produce this level of control. The viability of the wheat grain was not affected by any of the insecticidal treatments, with 90% germination recorded in treated and control, grains at all intervals tested.     

The inhibition of chitin synthesis in spodoptera littoralis larvae by flufenoxuron,teflubenzuron and diflubenzuron

The chitin precursor [¹⁴C] N-acetylglucosamine injected into the haemolymph of Spodoptera littoralis (Boisduval) larva was incorporated into the chitin exponentially with time. When caterpillars were injected with precursor at the commencement of feeding on acylurea-treated leaf discs, flufenoxuron, teflubenzuron and diflubenzuron were found to be equally effective inhibitors of chitin synthesis, measured after 21 h. The dose response curves by feeding are not parallel, indicating that the relative potency of the compounds will vary across the dose range. When chitin precursor was injected simultaneously with topically applied diflubenzuron, flufenoxuron or teflubenzuron, all three acylureas were found to be equally effective as inhibitors of chitin synthesis when measured after five hours. The I₅₀values (50% inhibition of chitin synthesis) were not significantly different; average 600 ng, compared with LD₅₀values (50% lethal dose) of ～ 13 ng for flufenoxuron and teflubenzuron but 130 ng for diflubenzuron (topical application). Injection of precursor 24 h after topical application of insecticide gave an I₅₀value which had dropped 670- and 150-fold for flufenoxuron and teflubenzuron respectively but only 20-fold for diflubenzuron. It is postulated that the reason for the low increase in diflubenzuron effectiveness with time was due either to less diflubenzuron than flufenoxuron reaching the site of action, or more probably, a faster rate of metabolism and excretion for diflubenzuron. The lower toxicity of diflubenzuron compared with flufenoxuron and teflubenzuron may not be due to any inherent differences in biochemical effectiveness, but rather to different penetration/metabolism properties.     

Egg-sterilizing effect of benzoylphenylureasvia the adult stage of the nitidulid beetleCarpophilus hemipterus

The effect of a 24-h exposure of adults ofCarpophilus hemipterus to artificial diets treated on the surface with different concentrations of benzoylphenylureas, on the hatchability of eggs laid during the subsequent 14 days, was investigated. The order of persistence of sterilization at 5 ppm was chlorfluazuron (IKI-7899), 12 days > XRD-473, 10 days > diflubenzuron, 6 days > teflubenzuron (CME 134), 4 days. Three of the substances (chlorfluazuron, diflubenzuron and teflubenzuron) had no direct ovicidal effect whenC. hemipterus eggs were dipped in 1000 ppm dilutions, but the larvae that hatched from the treatments died within 2 days.     

Effect of insecticide—synergist combinations on the survival of Spodoptera exigua

The chitin synthesis inhibitors diflubenzuron and teflubenzuron have recently become ineffective for the control of Spodoptera exigua in floricultural crops. An extended laboratory test with second-instar larvae of S. exigua on Vicia faba plants was carried out to determine the influence of synergists on the biological activity of three benzoylphenyl ureas (BPUs). The co-application of piperonyl butoxide, an oxidase inhibitor, did not increase the activity of diflubenzuron, teflubenzuron or hexaflumuron. The best results were obtained with diethyl maleate, for suppressing glutathione S-transferase activity, and with dimethoate, as a hydrolase inhibitor. A joint application of diflubenzuron (at a concentration which resulted in 43% survival) with diethyl maleate or dimethoate gave only 6.2 and 8.9% surviving larvae, respectively. In addition, development to fourth-instar larvae was inhibited. The more stable teflubenzuron was synergized by both compounds to a much lesser extent than diflubenzuron. None of the synergists had a significant effect on the activity of hexaflumuron, which was the most potent insecticide of the three BPUs tested against S. exigua.     

Synergism of teflubenzuron and chlorfluazuron in an acylurea-resistant field strain of Plutella xylostella L. (lepidoptera: Yponomeutidae)

Topical application of the synergists piperonyl butoxide (PB) and S,S,S-tributyl phosphorotrithioate (DEF) to second-instar larvae of a standard laboratory strain (FS) and an unselected Malaysian field strain (CH) of the diamondback moth Plutella xylostella had no significant effect on the toxicity of the acylurea insecticides, chlorfluazuron and teflubenzuron, in a subsequent leafdip bioassay. In contrast, pre-treatment with PB or DEF in acylurea-selected subpopulations of the CH strain with varying levels of cross-resistance to chlorfluazuron and teflubenzuron significantly increased (up to 34-fold and 28-fold, respectively) the toxicity of both compounds, suggesting that microsomal monooxygenases and esterases may be involved in resistance. The addition of a mineral oil, ‘Sunspray 6E’, to topically-applied chlorfluazuron consistently reduced its LD₅₀ value, and the effect of the oil appeared to be greatest on the most resistant population of P. xylostella. However, the effects of the oil were not significant (P > 0·05) and further studies are necessary to determine whether a penetration factor is present in the CH strain.     

Comparative laboratory evaluation of the acute and chronic toxicology of diflubenzuron,hexaflumuron and teflubenzuron against II instar desert locust (Schistocerca gregaria) (Orthoptera: Acrididae)

The acute and chronic toxicities of three benzoylphenyl ureas, diflubenzuron, hexaflumuron and teflubenzuron were assessed under laboratory conditions against two-day-old second (II) instar Desert Locust, Schistocerca gregaria (ForskÅl) nymphs. Following exposure by ingestion of a single precise dose applied to short pieces of spring barley, nymphs were monitored for two moults until the fourth (IV) instar. Analysis of acute response data gave three significantly different LD₅₀ statistics (P < 0·05), 68·0, 26·6 and 0·71 μg per nymph respectively for diflubenzuron, hexaflumuron and teflubenzuron. The probit regression slopes also differed significantly, indicating distinct tolerance distributions for the three compounds, the narrowest response being to diflubenzuron and the widest range of response being to teflubenzuron. The timing of death was found to vary between the compounds; most nymphs died during the first moult following treatment with either hexaflumuron or teflubenzuron. However, the majority of nymphs that died after exposure to diflubenzuron did so after completing the first moult after treatment, but before the second. The mean development times of nymphs during the II and especially third (III) instars were significantly longer (P < 0·05) than those of the controls following exposure to diflubenzuron and hexaflumuron. Teflubenzuron had no significant effect (P < 0·05) on the duration of the II instar. The potential of the three compounds to control S. gregaria populations in the field is discussed with particular reference to the timing and nature of acute and chronic responses.     

Genetic basis of resistance and studies on cross-resistance in a population of diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae)

The genetic basis of abamectin resistance was studied in a strain of the diamondback moth, Plutella xylostella (L), following laboratory selection of a field population collected at Xuanhua, Hebei Province, China. Data from the testing of F1 progeny from reciprocal crosses between abamectin-resistant and abamectin-susceptible strains indicated that resistance might be autosomal and incompletely recessive with a degree of dominance of -0.13. Chi-squared analyses from the response of a backcross of crossed F1 progeny and the resistant strain and F2 progeny were highly significant, suggesting that the resistance was probably controlled by more than one gene. The results of cross-resistance studies showed that there was little cross-resistance between abamectin and four pyrethroid insecticides (deltamethrin, beta-cypermethrin, fenvalerate and bifenthrin) and no cross-resistance between abamectin and the acylureas chlorfluazuron or flufenoxuron.     

小菜蛾对阿维菌素的抗性机制及交互抗性研究

用叶片药膜法研究了阿维菌素抗性小菜蛾 Plutella xylostella （L.）品系 对常用药剂的交互抗性谱以及增效醚（PB）和磷酸三苯酯（TPP）的增效作用。小菜蛾对阿 维菌素与高效氯氰菊酯、溴氰菊酯、氰戊菊酯和联苯菊酯等菊酯类药剂间具有比较低的交互 抗性，对后者抗性为3～20倍，对阿维菌素的抗性为575.6倍；对氟虫脲和氟啶脲没有交互抗 性。PB和TPP对阿维菌素分别增效8.2和5.5倍，说明小菜蛾对阿维菌素的抗性可能与多功能 氧化酶（MFO）和羧酸酯酶有关。通过差光谱技术测定了阿维菌素抗性和敏感小菜蛾细胞色 素P450的含量，抗性品系是敏感品系的1.38倍。     

Evidence for Resistance to Bacillus thuringiensis (Bt) subsp. kurstaki HD-1, Bt subsp. aizawai and Abamectin in Field Populations of Plutella xylostella from Malaysia

The efficacy of Bacillus thuringiensis (Bt) subsp. kurstaki HD-1 (‘Dipel’™; Btk; CryIA & CryII) and Bt. subsp. aizawai (‘Florbac’™; Bta; CryIA & CryIC) was assessed against larvae from various field populations of Plutella xylostella (F2 generation) collected in the Cameron Highlands, Malaysia in April 1994 and a lowland population (SERD 2; F10 generation) collected in December 1993. Evidence of resistance to Btk and to a lesser extent Bta is reported in these populations (LC₅₀ Toxicity Ratios [TR]=3–14 and 2–8 respectively), most notably in SERD 2. The first recorded evidence of resistance to abamectin (TR=17–195-fold) in field populations of P. xylostella is also reported. In an unselected sub-population of SERD 2, the TR values for Btk, Bta and abamectin declined 2- to 3-fold (P<0·01) over six generations in the laboratory (F10–F16) while in sub-populations of SERD 2 selected with these products (F11–F15) there was a significant (P<0·01) increase in the TR (15-, 3- and 2·5-fold respectively) when compared with the F10 generation. This suggests the presence of marked resistance to Btk and some resistance to Bta and abamectin. There is also evidence of slight cross-resistance to Btk in the Bta-selected sub-population but no evidence for the reverse selection of resistance or for cross-resistance between Btk and abamectin. Concurrent selection studies (F11–F15) with another sub-population of SERD 2 demonstrated resistance to the acylurea insect growth regulator, teflubenzuron (‘Nomolt’™) (29-fold increase in TR). Based on the selection experiments with SERD 2, estimates of realised heritability (h²) of resistance gave very high values for teflubenzuron and Btk (c.0·7) and moderate values for abamectin and Bta (c.0·3). The results are discussed in relation to integrated pest management (IPM) and insecticide resistance management (IRM) strategies for P. xylostella.     

Laboratory selected resistance to diflubenzuron in larvae of aedes aegypti

The variation in tolerance to diflubenzuron [1-(4-chlorophenyl)-3-(2,6-difluorobenzoyl)urea] was examined in fourth instar larvae of seven strains of Aedes aegypti, some of which were resistant to DDT and permethrin. The difference between the least and the most tolerant to diflubenzuron was approximately two-fold. There was no correlation with resistance to the other insecticides. A DDT-resistant strain (T8) was selected 10 times (during 12 generations) with diflubenzuron. The LC₅₀ to diflubenzuron had increased 3.3 times by the S₈ generation but there was no further increase in later generations despite further selection. Associated with this increase, a marked decrease in resistance to DDT was observed but no change in permethrin tolerance. A genetically enriched strain (Hotchpotch) was synthesised from 35 strains of different geographic origin and crossed to the selected T8 strain before subsequent generations were selected five times with diflubenzuron. This procedure resulted in an 8 to 12-fold increase in the LC₅₀ value over that for unselected T8, accompanied by a decrease in the slope of the log dose against probit mortality line.     

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.     

Rapid report acetamiprid resistance and cross-resistance in the diamondback moth, Plutella xylostella

A 110-fold acetamiprid-resistant Plutella xylostella (L) strain was established after four selection experiments (in five generations) on a 9.5-fold resistant colony in the laboratory. The resistant strain did not show cross-resistance to chlorfluazuron or Bacillus thuringiensis subsp kurstaki Berliner, but displayed low resistance to cartap and phenthoate.     

甜菜夜蛾对虫酰肼的抗药性研究初报

在室内用虫酰肼对甜菜夜蛾幼虫进行抗药性选育,经过12代汰选,甜菜夜蛾对虫酰肼产生了5.47倍的抗性,抗性发展缓慢。该种群对甲氧虫酰肼有一定交互抗性,对有机磷、氨基甲酸酯、拟除虫菊酯类等多种杀虫剂都不存在交互抗性,抗性比值在0.83~1.63倍之间。     

广州、深圳地区小菜蛾对定虫隆、Bt的抗性监测

采用 Potter's 喷雾塔喷雾的方法测定广州和深圳菜区小菜蛾对儿丁质合成抑制剂定虫隆、氟虫脲、伏虫隆、氟铃脲的抗药性。结果表明,深圳菜区小菜蛾对定虫隆产生明显抗性,1990年11月测定抗性倍数为15.0,1991年12月为70.9,对氟虫脲、伏虫隆、氟铃脲具有不同程度交互抗性,抗性倍数分别为26.7,15.5 和5.8。广州菜区小菜蛾对以上药剂均未产生抗性(抗性倍数最高为2.8)。采用浸渍法测定小菜蛾对苏云金杆菌 Bt 抗性的结果表明,深圳菜区小菜蛾对 Bt 的抗性逐年提高,1990年12月测定的抗性倍数为21.1,1991年12月为26.9,1992年3月为35.0,广州菜区小菜蛾具有强的耐药性或低抗药性,抗性倍数8—9。     

Cross-resistance and biochemical mechanisms of abamectin resistance in the western flower thrips, Frankliniella occidentalis

Abamectin resistance was selected in the western flower thrips [Frankliniella occidentalis (Pergande)] under the laboratory conditions, and cross-resistance patterns and possible resistance mechanisms in the abamectin-resistant strain (ABA-R) were investigated. Compared with the susceptible strain (ABA-S), the ABA-R strain displayed 45.5-fold resistance to abamectin after 15 selection cycles during 18 generations. Rapid reversion of abamectin resistance was observed in the ABA-R strain in the absence of the insecticide selection pressure. Moderate and low levels of cross-resistance to chlorpyrifos (RR 11.4) and lambda-cyhalothrin (3.98) were observed in the ABA-R strain, but no significant cross-resistance was found to spinosad (2.00), acetamiprid (1.47) and chlorfenapyr (0.26). Our studies also showed that the esterase inhibitor S,S,S-tributyl phosphorotrithioate (DEF) and glutathione S-transferase inhibitor diethyl maleate (DEM) were not able to synergize the toxicity of abamectin, whereas the oxidase inhibitor piperonyl butoxide (PBO) conferred a significant synergism on abamectin in the ABA-R strain (SR 3.00). Biochemical analysis showed that cytochrome P450 monooxygenase activity of the ABA-R strain was 6.66-fold higher than that of the ABA-S strain. It appears that enhanced oxidative metabolism mediated by cytochrome P450 monooxygenases was a major mechanism for abamectin resistance in the western flower thrips.     

Effect of chitin-synthesis inhibitors on stored-product beetles

The fecundity of four important coleopteran stored-grain pests, Rhyzopertha dominica (Fab.), Sitophilus oryzae (L.), Oryzaephilus surinamensis (L.) and Tribolium castaneum (Herbst.) was assessed after exposure to wheat treated with five benzoylphenyl ureas (BPUs), members of the chitin-synthesis inhibitor group of insect growth regulators (IGRs). Chlorfluazuron was found to control all four species more effectively than triflumuron, teflubenzuron, flufenoxuron or diflubenzuron. Two weeks of exposure of parents to 0.5 mg kg ?1 of all the BPUs, except diflubenzuron, inhibited almost all F1 production and all the F2 production of each species. The fecundities of T. castaneum and O. surinamensis recovered almost to their untreated levels after two weeks of exposure to 1 mg kg ?1 of BPUs followed by two weeks on untreated wheat, but the effect of the exposure persisted for at least two weeks on untreated wheat in adult S. oryzae and to a lesser extent in R. dominica. The mean F1 response of the four species after two weeks of exposure to four doses showed clearly that the four newer BPUs were all similar in their overall effectiveness and significantly more effective than diflubenzuron, although diflubenzuron was equally effective against S. oryzae. At 0.1 mg kg ?1 triflumuron was less effective against O. surinamensis, while teflubenzuron and flufenoxuron were not very effective against the rice weevil, S. oryzae. Chlorfluazuron provided the most effective control of all four stored-product pest species, including the weevil, S. oryzae. The BPUs show particular potential for use in the stored-product industry because low dosages will effectively control the rice weevil in addition to the other major pest species.     

Indoxacarb resistance in the house fly, Musca domestica

Indoxacarb (DPX-MP062) is a recently introduced oxadiazine insecticide with activity against a wide range of pests, including house flies. It is metabolically decarbomethoxylated to DCJW. Selection of field collected house flies with indoxacarb produced a New York indoxacarb-resistant (NYINDR) strain with >118-fold resistance after three generations. Resistance in NYINDR could be partially overcome with the P450 inhibitor piperonyl butoxide (PBO), but the synergists diethyl maleate and S,S,S-tributyl phosphorothioate did not alter expression of the resistance, suggesting P450 monooxygenases, but not esterases or glutathione S-transferases are involved in the indoxacarb resistance. Conversely, the NYINDR strain showed only 3.2-fold resistance to DCJW, and this resistance could be suppressed with PBO. Only limited levels of cross-resistance were detected to pyrethroid, organophosphate, carbamate or chlorinated hydrocarbon insecticides in NYINDR. Indoxacarb resistance in the NYINDR strain was inherited primarily as a completely recessive trait. Analysis of the phenotypes vs. mortality data revealed that the major factor for indoxacarb resistance is located on autosome 4 with a minor factor on autosome 3. It appears these genes have not previously been associated with insecticide resistance.     

Mechanisms of resistance to diflubenzuron in the house fly, Musca domestica (L.)

The mechanisms of resistance to the chitin synthesis inhibitor diflubenzuron were investigated in a diflubenzuron-selected strain of the house fly (Musca domestica L.) with > 1000 × resistance, and in an OMS-12-selected strain [O-ethyl O-(2,4-dichlorophenyl)phosphoramidothioate] with 380 × resistance to diflubenzuron. In agreement with the accepted mode of action of diflubenzuron, chitin synthesis was reduced less in larvae of the resistant (R) than of a susceptible (S) strain. Cuticular penetration of diflubenzuron into larvae of the R strains was about half that of the S. Both piperonyl butoxide and sesamex synergized diflubenzuron markedly in the R strains, indicating that mixed-function oxidase enzymes play a major role in resistance. Limited synergism by DEF (S,S,S-tributyl phosphorotrithioate) and diethylmaleate indicated that esterases and glutathione-dependent transferases play a relatively small role in resistance. Larvae of the S and R strains exhibited a similar pattern of in vivo cleavage of ³H- and ¹⁴C-labeled diflubenzuron at N₁C₂ and N₁C₁ bonds. However, there were marked differences in the amounts of major metabolites produced: R larvae metabolized diflubenzuron at considerably higher rates, resulting in 18-fold lower accumulation of unmetabolized diflubenzuron by comparison with S larvae. Polar metabolites were excreted at a 2-fold higher rate by R larvae. The high levels of resistance to diflubenzuron in R-Diflubenzuron and R-OMS-12 larvae are due to the combined effect of reduced cuticular penetration, increased metabolism, and rapid excretion of the chemical.