Comparative metabolism of 1,1-Bis-(p-chlorophenyl)-2-nitropropane (Prolan) in mouse,insects, and in a model ecosystem

¹⁴C-labeled Prolan or 1,1-bis-(p-chlorophenyl)-2-nitropropane was found to be some-what more biodegradable than DDT. This insecticide, although highly resistant to microsomal metabolism, was degraded by elimination to 1,1-bis-(p-chlorophenyl)-1-propene, and by reduction to 1,1-bis-(p-chlorophenyl)-2-aminopropane. The major degradative pathway, however, was by oxidation to 1,1-bis-(p-chlorophenyl)-2-propanone, to 1,1-bis-(p-chlorophenyl)-pyruvic acid, to bis-(p-chlorophenyl)-acetic acid, and ultimately to p,p′-dichlorobenzophenone. Therefore the ultimate degradative products of Prolan are identical to those produced from DDT.     

An evaluation of the persistence in soil of two non-chlorinated insecticides analogous to DDT

Two insecticides 1,1-bis(4-ethoxyphenyl)-2-nitrobutane (ENB) and 1,1-bis(4-ethoxyphenyl)-2-nitropropane (ENP), at concentrations varying from 0.4-40 mg kg^?1 have been examined for persistence in three soils. The proportions of the original compounds recovered decreased rapidly in a clay loam (falling to 50% in 2-6 weeks), decreased slowly in a sand (falling to 50% in 16-64 weeks) and decreased variably in a heavy clay (falling to 50% in 2-10 weeks). After the initial phase, the proportion recovered generally decreased at a slower rate. Breakdown of ENP appeared to proceed via a stepwise oxidation to give 1,1-bis(4-ethoxyphenyl)propan-2-one and 4,4′-diethoxybenzophenone. These products were readily broken down under aerobic conditions and were isolated only from waterlogged or sterile soil. Experiments under sterile conditions indicated that biological breakdown was a major factor and that chemical breakdown was specifically surface-activated.     

Diversity and frequencies of genetic mutations involved in insecticide resistance in field populations of the house fly (Musca domestica L.) from China

Insecticides have been extensively used for house fly control in China, with dichlorvos and deltamethrin being widely used. Knowledge about the current status of insecticide resistance and the underlying genetic changes is crucial for developing effective fly control strategies. The susceptibility to dichlorvos and deltamethrin, and the frequencies of genetic mutations involved in insecticide resistance were studied in five field populations of the house fly collected across China. Bioassay results show that flies exhibit 14- to 28-fold resistance to dichlorvos and 41- to 94-fold resistance to deltamethrin, indicating that dichlorvos and deltamethrin resistance are common in house fly populations in China. Molecular analysis reveals that flies from the five various locations carry resistance alleles at multiple loci and have diverse allelic types, different relative frequencies and combinations of each allele. Four non-synonymous single nucleotide polymorphisms (SNPs) (i.e. V260L, G342A/V, F407Y) in acetylcholinesterase (Ace) and two mutations (W251L/S) in a carboxylesterase (MdαE7) were commonly present in the field house flies. The L1014H rather than L1014F mutation in the voltage sensitive sodium channel gene (Vssc) was widely distributed in Chinese house flies. CYP6D1v1, which confers pyrethroid resistance, was found in all the five tested populations in China, although its frequency in house fly from Shandong province was very low. Our results suggest that resistance monitoring and management of house flies should be customized for a given location.     

Decreased nerve sensitivity and decreased cuticular penetration as mechanisms of resistance to pyrethroids in a (1R)-trans-permethrin-selected strain of the house fly

A fenthion-resistant strain of the house fly (Musca domestica L.) was selected with bioresmethrin resulting in ca. 90-fold resistance to the selecting agent. This strain was subsequently selected with (1R)-trans-permethrin producing ca. 140-fold resistance to this latter insecticide. The permethrin-resistant (147-R) strain was highly cross-resistant to several other pyrethroids and demonstrated resistance to knockdown by these insecticides as well as by DDT. The sensitivity of the central nervous system to four pyrethroids was investigated. The 147-R strain was 2.6-fold less sensitive to (1R)-trans-ethanoresmethrin than the susceptible (NAIDM-S) strain, and >43-fold and >67-fold less sensitive to (1R,S)-cis, trans-tetramethrin and (1R)-trans-permethrin, respectively. It also displayed decreased penetration of (1R,S)-trans-[¹⁴C]permethrin when compared to the NAIDM-S strain. Lower nerve sensitivity and decreased cuticular penetration are potential mechanisms of resistance to pyrethroids in house flies in the United States.     

Structure-activity correlations of the insecticide Prolan and its analogs

Structure-activity correlations for 45 insecticidal diaryl nitropropanes (Prolan analogs) were analyzed by multiple regression analysis. Molecular bulk constants including van der Waal's radii, molar attraction constants, parachor, steric constants such as Taft's E₈ and Verloop's dimensional steric constants, hydrophobic constants such as II, and electronic parameters such as σ, F, and R were evaluated. It was concluded that the diaryl nitropropanes like the diaryl trichloroethanes fit into a receptor site which has an optimum volume for maximum interaction. The interaction between the insecticide and the receptor shows high correlation with steric constants for the aryl substituents and with intermolecular attractive forces. Highly asymmetrical compounds such as 1-(p-fluorophenyl)-1-(p-hexoxyphenyl)-2-nitropropane were surprisingly effective insecticides.     

Fitness cost, cross resistance and realized heritability of resistance to imidacloprid in Spodoptera litura (Lepidoptera: Noctuidae)

Imidacloprid has been used as a key insecticide for controlling sucking insect pests of cotton, whereas Spodoptera litura also has been indirectly exposed to this insecticide in Pakistan. To evaluate the risk of resistance evolution and to develop a better resistance management strategy, a field collected population was selected with imidacloprid in the laboratory. Thereafter, fitness cost, realized heritability and cross resistance of imidacloprid resistance in S. litura were investigated. After 14 generations of selection with imidacloprid, S. litura developed a 137.48-fold resistance to the insecticide. Bioassay revealed that this strain showed cross-resistance to acetamiprid (RR 8.52) and a little to lamdacyhalothrin (1.92) but negative cross-resistance was found to methomyl (−0.19). The resistant strain had a relative fitness of 0.38, with substantially lower rates of larval survival, larval duration, male pupal duration, development time, emergence rate of healthy adults, fecundity, hatchability, and prolonged larval and pupal duration. Mean relative growth rate of the larvae, intrinsic rate of population increase, and biotic potential was lower for the selected populations. The estimated realized heritability (h²) of imidacloprid resistance was 0.15 in the resistant strain of S. litura. Development of the resistance may cost significant fitness for the resistant population. This study provided valuable information for further understanding the impact of imidacloprid resistance on physiological parameters of S. litura and for facilitating the development of resistance management strategies.     

Effect of dihydrodillapiole on pyrethroid resistance associated esterase inhibition in an Indian population of Spodoptera litura (Fabricius)

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.     

The A302S mutation in Rdl that confers resistance to cyclodienes and limited cross-resistance to fipronil is undetectable in field populations of house flies from the USA

Fipronil is a relatively new insecticide with great potential for insect control, however widespread use of cyclodiene insecticides has selected for an A302S mutation in the Rdl (GABA gated chloride channel) allele. This mutation gives resistance to cyclodienes and limited cross-resistance to fipronil. Given the concern over the possible reduction in efficacy and/or lifetime that fipronil might be used for pest control (given the extensive use of cyclodienes in the past), it is important to know the frequency of the A302S Rdl mutation in field populations. To ascertain the relative frequency of the A302S Rdl mutation in house fly populations we used three experimental approaches. First, we attempted to select for fipronil resistance by initially treating 33,100 field collected flies and selecting 14 additional generations. We were unable to produce a highly resistant strain. Second, we directly sequenced field collected flies. Third, we tested field collected house flies with a diagnostic dose of dieldrin and then genotyped the survivors. Out of the 4750 flies tested, there were no Rdl resistance alleles detected. We conclude that the resistant Rdl allele is rare in house flies in the US due to decades without cyclodiene use and a fitness disadvantage (in the absence of cyclodienes) of the 302S Rdl allele. The limited cross-resistance provided by the cyclodiene resistant Rdl allele, combined with the very low frequency of this allele in field populations, suggests that fipronil could be a promising insecticide for house fly control.     

In vitro inhibition of acetylcholinesterase by O,O-dimethyl S-aryl phosphorothioates

In vitro inhibition of house cricket head, house fly head, and bovine erythrocyte acetylcholinesterase by O,O-dimethyl S-aryl phosphorothioates was studied by Main's kinetic treatment. The potency of the compounds as reflected by the bimolecular reaction constants (k_i) indicated that house fly head acetylcholinesterase was the most sensitive to the inhibition followed by house cricket head and bovine erythrocyte acetylcholinesterase. There are no linear relationships between the phosphorylation rate constants and the total binding energies for the inhibition of three enzymes by this series of compounds, suggesting that the initial binding and the phosphorylation rate are not related. The structure and activity relationships were analyzed by multiple regression analyses with the use of Hammett's sigma, alkaline hydrolysis rates of the compounds, and p_i constants. The hydrophobic bonding of the compound on the enzyme surface as reflected by the p_i constant played a significant role in the determination of the potency of the inhibition of house cricket head and house fly head acetylcholinesterase by those compounds. However, the alkaline hydrolysis rates of the compounds, or the Hammett's sigma values seems to play a more important role in the determination of the inhibition of bovine erythrocyte acetylcholinesterase. Moderate insecticidal activity toward house crickets, house flies, and mosquito larvae were found.     

Cytochrome P-450 in insects: 2. Multiple forms in the flesh fly (Sarcophaga bullata,Parker), and the blow fly (Phormia regina (Meigen))

Microsomes prepared from the abdomens of the flesh fly (Sarcophaga bullata, Parker) and the blow fly (Phormia regina (Meigen)) contain approximately one-fifth and one-eighth as much cytochrome P-450, respectively, as those prepared from house fly (Musca domestica, L.) abdomens. These values correlate well with the microsomal aldrin epoxidase activity of the three species and with their respective susceptibilities to the insecticide, propoxur. When the microsomes of the flesh fly and the blow fly are solubilized by treatment with deoxycholate and resolved by ion-exchange chromatography on DEAE-cellulose and hydroxylapatite, four chromatographically distinct fractions containing cytochrome P-450 are obtained. Spectrophotometric assays of the cytochrome P-450 in these fractions indicate purifications of two-to sixfold for the flesh fly hemoprotein and two-to eightfold for that of the blow fly. SDS-Polyacrylamide gel electrophoresis of the four column fractions from the flesh fly microsomes indicates that six hemoproteins in the 40,000–60,000 molecular weight range are present. In similar experiments with blow fly fractions containing approximately the same amount of cytochrome P-450 no high molecular weight hemoproteins could be detected. This result is interpreted, with other evidence, as an indication of the greater instability of the blow fly hemoprotein. The results indicate that multiple forms of cytochrome P-450 are present in both species but there is insufficient data on which to estimate the number of such forms.     

新型杀虫剂甲磺虫腙的合成与杀虫活性

以对氯苯甲酰氯、苯甲醚、甲基磺酰氯、水合肼、丁酮等为原料合成了新型杀虫剂甲磺虫腙(试验号:ZJ0967),其化学名称为甲磺酸-4- 苯酯,其结构经1H NMR、MS和元素分析确认。室内生测结果表明,甲磺虫腙对小菜蛾Plutella xylostella、甜菜夜蛾Spodoptera exigua,特别是对粘虫Mythimna sepatara、斜纹夜蛾Prodenia litura表现出良好的杀虫活性。进一步毒力测定表明,甲磺虫腙对斜纹夜蛾的LC50值为8.45(7.69~9.62) mg/L,其杀虫活性与虫酰肼相当,但低于氟啶脲。田间试验结果表明,70%甲磺虫腙WG在有效成分为210~420 g/hm2时,对斜纹夜蛾7 d的防效可达88.6% ~93.9%,未观察到对作物产生药害。     

Insecticide resistance in house flies from the United States: Resistance levels and frequency of pyrethroid resistance alleles

Although insecticide resistance is a widespread problem for most insect pests, frequently the assessment of resistance occurs over a limited geographic range. Herein, we report the first widespread survey of insecticide resistance in the USA ever undertaken for the house fly, Musca domestica, a major pest in animal production facilities. The levels of resistance to six different insecticides were determined (using discriminating concentration bioassays) in 10 collections of house flies from dairies in nine different states. In addition, the frequencies of Vssc and CYP6D1 alleles that confer resistance to pyrethroid insecticides were determined for each fly population. Levels of resistance to the six insecticides varied among states and insecticides. Resistance to permethrin was highest overall and most consistent across the states. Resistance to methomyl was relatively consistent, with 65–91% survival in nine of the ten collections. In contrast, resistance to cyfluthrin and pyrethrins + piperonyl butoxide varied considerably (2.9–76% survival). Resistance to imidacloprid was overall modest and showed no signs of increasing relative to collections made in 2004, despite increasing use of this insecticide. The frequency of Vssc alleles that confer pyrethroid resistance was variable between locations. The highest frequencies of kdr, kdr-his and super-kdr were found in Minnesota, North Carolina and Kansas, respectively. In contrast, the New Mexico population had the highest frequency (0.67) of the susceptible allele. The implications of these results to resistance management and to the understanding of the evolution of insecticide resistance are discussed.     

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.     

Insecticide metabolism by multiple glutathione S-transferases in two strains of the house fly,Musca domestica (L)

Glutathione S-transferases from two strains of house fly have been prepared in a high degree of homogeneity by a procedure involving affinity chromatography and isoelectrofocusing. They fall into two groups in each strain. One group, of isoelectric point greater than pH 6.5, catalyzes the glutathione-dependent degradation of lindane, diazinon and methyl parathion. The other group, of low isoelectric point, has conjugating activity with the model substrate CDNB, but very little activity with the insecticide substrates. In the Cornell R strain the three isoenzyme forms in the high pI group appear to be almost identical in their substrate specificities. In the A strain, it is apparent that the enzyme forms falling into this group vary markedly in substrate specificity. The dehydrochlorination of DDT paralleled very closely the conjugation of the other insecticides catalyzed by the three high pI enzymes in the Cornell R strain. In the A strain, DDT dehydrochlorinase was most strongly associated with a glutathione S-transferase isoelectric at pH 7.1. It is tentatively concluded that multiple genes are involved in the production of the glutathione S-transferases involved in pesticide metabolism in the house fly and that DDT dehydrochlorinase may be derived from some, but not all, of these same genes.     

Structural modifications increase the insecticidal activity of ryanodine

The toxicity of ryanodine ( 1 ) and 9,21-didehydroryanodine ( 2 ) (the principal active ingredients of the botanical insecticide ryania) to adult female house flies (Musca domestica L.) is attributable to binding to the ryanodine receptor (ryr) and thereby disrupting the Ca²⁺-release channel. These ryanoids, assayed in house flies with piperonyl butoxide (PBO) to suppress cytochrome P450-dependent detoxification, give injected KD₅₀ values of 0·07–0·11 μg g^-1, injected LD₅₀ values of 0·39–0·45 μg g^-1 and topical LD₅₀ values of 12– 50 μg g^-1. They inhibit the [³H]ryanodine binding site of house fly and rabbit muscle with IC₅₀ values of 3–10 nM . This study examines the effect of structure on potency, with 15 variants of the cyclohexane substituents, two 4,6-cyclic boron and two methylated derivatives, and four modifications of the isopropyl and ester substituents. The most effective compound examined was 10-deoxy- 2 ( 3 ) which was more potent than 2 by 2–4-fold on injection and 29-fold applied topically following PBO (LD₅₀ 0·41 μg g^-1). Additional high-potency compounds were 10-oxo- 1 and the cyclohexane variants with lactam, 21-nor-9-oxo and 21-nor-10-deoxy substituents. Other modifications usually reduced toxicity. The injected knockdown potency of the ester ryanoids was generally related to their effectiveness in competing with [³H]ryanodine at the ryr of rabbit skeletal muscle. Two non-ester ryanoids, ryanodol and 9,21-didehydroryanodol, were found to be more toxic than predicted from their potency at the ryr and may therefore act in a different manner such as at a K⁺ channel, as suggested by Usherwood and Vais. Clearly ryanoids are challenging prototypes for a potential new generation of insecticides. © 1997 SCI.     

Cytochrome P-450 in insects: 4. Reconstitution of cytochrome P-450-dependent monooxygenase activity in the house fly

Two cytochrome P-450-containing fractions were isolated from detergent-solubilized house fly microsomes by hydrophobic chromatography on a tryptamine-Sepharose gel. These fractions (designated P-450-1 and P-450-2) were distinctive in their spectral characteristics and in their profiles following electrophoresis in the presence of sodium dodecyl sulfate. Both fractions exhibited NADPH-dependent epoxidase activity when reconstituted with purified house fly cytochrome P-450 reductase and phospholipid. The aldrin epoxidase activity of fraction P-450-1 was twice that of P-450-2 even though heptachlor epoxidase activity of the fractions was equivalent. O-Demethylase activity with 7-methoxy-4-methylcoumarin was detectable only in the P-450-2 fraction.     

Protein metabolism in Spodoptera litura (F.) is influenced by the botanical insecticide azadirachtin

Azadirachtin, as a botanical insecticide, affects a wide variety of biological processes, including reduction of feeding, suspension of molting, death of larvae and pupae, and sterility of emerged adults in a dose-dependent manner. However, the mode of action of this toxin remains obscure. By using proteomic techniques, we analyzed changes in protein metabolism of Spodoptera litura (F.) induced by azadirachtin. Following feeding 4th instar larvae of Spodoptera litura (F.) with an artificial diet containing 1 ppm azadirachtin until pupation, 48 h old pupae were collected and protein samples prepared. Total soluble protein content was measured and the results showed that azadirachtin significantly influenced protein level. Moreover, the proteins were separated by 2-DE (two-dimensional polyacrylamide gel electrophoresis) and 10 proteins were significantly affected by azadirachtin treatment when compared to an untreated control. Six of these proteins were identified with peptide mass fingerprinting using MALDI-TOF-MS after in-gel trypsin digestion. These proteins are involved in various cellular functions. One identified protein may function as an ecdysone receptor, which regulates insect development, and reproduction. It is suggested that the botanical insecticide azadirachtin affects protein expression and the azadirachtin-related proteins would be essential for a better understanding of the mechanisms by which neem toxins exert their effects on insects.     

Penetration and metabolism of 2-isopropoxyphenyl N-methyl-N-(2-methyl-4-tert-butylphenylsulfenyl) carbamate in the house fly and honeybee

The penetration and metabolism of 2 - isopropoxyphenyl N-methyl-N-(2-methyl-4-tert - butylphenylsulfenyl)carbamate or sulfenyl-propoxur was examined in the house fly and honeybee. Reduced penetration was found to be a factor contributing to the lower toxicity of sulfenyl-propoxur to the honeybee. Honeybees and house flies metabolized sulfenyl-propoxur qualitatively in a similar manner. Quantitatively, larger amounts of propoxur were found in the house fly than in the honeybee soon after treatment with sulfenyl-propoxur. The slower rate of conversion of sulfenyl-propoxur to propoxur was considered as another factor responsible for the lower toxicity of the sulfenylated derivative to bees. The high susceptibility of bees to propoxur was related to high internal amounts of unchanged propoxur found soon after treatment.     

Characterization of multiple glutathione transferases from the house fly, Musca domestica (L)

Glutathione transferases have been purified to a high degree of homogeneity from three strains of house fly by a procedure involving affinity chromatography on glutathione-sulfobromophthalein conjugate immobilized on Sepharose 4B, followed by preparative isoelectrofocusing. The affinity chromatography yielded purifications of between about 10- and 100-fold, depending on the strain and the substrate with which activity was measured. Each strain was shown to possess several proteins with glutathione S-transferase activity which fell into two clearly defined groups. The first group, of relatively low isoelectric point, showed activity with CDNB but little with DCNB, p-nitrobenzylchloride, or 1,2-epoxy-3-(p-nitrophenoxy)propane, whereas the second group, of higher isoelectric points, showed substantial activity with all substrates tested. Studies on the subunit structure of these enzymes demonstrated the existence of three different sized subunits of M_r 20,000, 22,000, and 23,500. From the experimental evidence recorded here, the existence of at least three functionally different glutathione transferases is inferred.