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A 3-yr field study was conducted at commercial grape (Vitis spp.) farms to evaluate insect management programs for control of the grape berry moth, Paralobesia viteana Clemens (Lepidoptera: Tortricidae) and conservation of natural enemies. At each farm, one vineyard received only reduced-risk insecticides for control of second and third generation P. viteana, whereas the comparison vineyard received conventional insecticides. Both vineyards received a conventional insecticide application for control of first generation P. viteana and other insect pests. Monitoring with pheromone traps showed no differences between programs in the total number of adult male moths trapped in vineyards, and oviposition by P. viteana was similar between the two programs in all 3 yr. During weekly samples of crop infestation, both programs had a similar percentage of clusters infested by P. viteana larvae. Berries infested by P. viteana were collected from vineyard borders during the second and third P. viteana generations and held under controlled conditions. In eight of the nine berry samples, survival of larvae was significantly lower in berries collected from vineyards managed under the reduced-risk insecticide program compared with the conventional program. Parasitism of P. viteana larvae in these samples was not consistently different between the two insecticide programs over 3 yr, and similar captures of natural enemies were found on yellow sticky traps in the two programs throughout the study. Our results indicate that integrated pest management programs incorporating reduced-risk insecticides for control of P. viteana can obtain similar or greater control of P. viteana compared with programs based solely on conventional insecticides, but they may not lead to measurable long-term increases in parasitism of P. viteana. [ABSTRACT FROM AUTHOR]

Susceptibility to methoxyfenozide of Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae) larvae was determined through exposure of neonate and fourth instars to dipped and sprayed pepper, Capsicum annum L., leaves. Methoxyfenozide and spinosad were tested against adults of this noctuid by oral, residual, and topical application. In larvae, we evaluated five (range, 0.001–10 mg active ingredient [Al]/liter) and six (range, 1–250 mg [AI]/liter) concentration levels by instars, respectively, by using two application methods at three different age leaf residues (0, 3, and 6 d after application). According to LC50 values, no significant differences were observed between the same age leaf residues of both application methods at 96 and 72 h after ingestion treatment on neonates and fourth instars, respectively. Nevertheless, toxicity of methoxyfenozide decreased significantly after time. For both application methods, the LC50 values of the first leaf residue (0 d after application) were significantly lower than those of 6-d-old residues. Furthermore, larval weight of fourth instars fed for 48 h with pepper, Capsicum annum L., leaves containing methoxyfenozide was significantly suppressed. Spinosad and methoxyfenozide reduced in a dose-dependent manner the fecundity and fertility of S. littoralis adults when treated oral and residually. Likewise, when methoxyfenozide was administered orally in three different adult crosses, the fecundity was strongly affected, independently of the treated sex. We conclude that the combination of lethal and sublethal effects of methoxyfenozide and spinosad might exhibit significant effects on the population dynamics of S. littolaris. [ABSTRACT FROM AUTHOR]

The chemical communication (female attractiveness and male responsiveness) of adult oriental fruit moth, Grapholita inolesta (Busck) (Lepidoptera: Tortricidae), exposed to surfaces treated with the ecdysteroid agonist methoxyfenozide for 48 h were investigated in two laboratory wind tunnel assays. The recapture assay examined the ability of treated males to orient to a single cage of treated females, and the data gathered were mean percentage of males recaptured per treatment. The male sexual behavior assay examined some specific orientation behaviors (associated with sexual excitability) of treated males when they were given a choice of two competing pheromone sources (cages of treated females), and the data gathered were mean time males spent in upwind plume orientations and at source contact (female cage) per treatment. Data from the recapture assay suggests that exposure to methoxyfenozide impacts male responsiveness more than female attractiveness. In contrast, data from the sexual behavior assay strongly revealed that exposure to methoxyfenozide-treated surfaces does negatively impact both the ability of calling females to attract males and of aroused males to display sustained upwind flight behavior and time spent at the female cages. [ABSTRACT FROM AUTHOR]

The toxicity of spinosad and methoxyfenozide against neonates and fourth instars of Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae) was tested under laboratory conditions. According to LC50 values, no significant differences were observed between spinosad (0.50 mg [AI]/kg diet) and methoxyfenozide (0.54 mg [AI]/kg diet) after 48 h of ingestion treatment on neonate larvae, based on the overlap of 95% CU Similarly, on fourth instars, no significant differences were observed between LC50 (2.98 and 5.17 mg [AI]/kg diet for spinosad and methoxyfenozide, respectively, at 96 h after ingestion of artificial diet) and LD50 (4.74 and 2.68 μg [AI]/g larva for spinosad and methoxyfenozide, respectively, at 144 h after topical application). In addition, spinosad and methoxyfenozide significantly suppressed weight gain of neonates and fourth instars continuously fed with artificial diet containing the insecticides. The second part of this project was focused on the effects of methoxyfenozide on the reproduction of S. littoralis by determining the effect on oviposition in three different crosses and the pharmacokinetics pattern of (14C)-methoxyfenozide in the body tissues of female and male adults after topical application in their excrement and in the deposited eggs to test transport through the female and the male adult. Methoxyfenozide negatively affected the reproduction of adults regardless the treated sex, presumably because of its presence in the adult body, accumulation in the laid eggs, and slow excretion of the product. We conclude that spinosad and methoxyfenozide represent an important choice to be used in integrated pest management where S. littoralis is a major pest. [ABSTRACT FROM AUTHOR]

This study examined the lethal and sublethal effects on the beneficial insect Bombus terrestris by two classes of insect growth regulators (IGRs) that are commercially used in agriculture to control pest insects. Three juvenile hormones analogues (JHAs) (pyriproxyfen, fenoxycarb and kinoprene) and two ecdysone agonists or moulting accelerating compounds (MACs) (tebufenozide and methoxyfenozide) were tested. The bumblebee workers were exposed to the insecticides via three different routes of exposure: dermally by topical contact, and orally via the drinking sugar water or the pollen. In the first series of experiments the IGRs were applied at their respective maximum field recommended concentration (MFRC). These risk hazard tests showed that the tested IGRs caused no acute toxicity on the workers, and any compound had an adverse effect on reproduction (production of males). In addition, larval development was followed in the treated nests compared with the controls. After application of the two MACs and the JHA fenoxycarb no adverse effects were observed on larval development. However, in the nests where the workers were exposed to the JHAs pyriproxyfen and kinoprene higher numbers of dead larvae were scored. These larvae were third and fourth instars, implying a lethal blockage of development before metamorphosis. In a second test, a series of dilutions was made for kinoprene, and these results revealed that only the MFRC caused a toxic effect on the larval development. On the other hand, kinoprene at lower concentrations (0.0650 mg ai/l) had a stimulatory effect on brood production. It was remarkable that ovaries of such treated dominant workers were longer and contained more eggs than in the controls. In a last experiment, the cuticular uptake was determined for a JHA and MAC to evaluate to what extent worker bees accumulate these classes of IGRs. Cuticular uptake ranged from 34 to 83% at 24 h after topical application. Overall, the obtained results indicate that the tested IGRs at their recommended concentration are safe to be used in combination with B. terrestris. [ABSTRACT FROM AUTHOR]

Nests of yellowjacket wasp, Vespula germanica F., were collected, from which comb pieces were selected with last stage larvae (n ≥ 35), that were later maintained at 25°C and ∼50% RH in absence of light. Laboratory assays were conducted to determine the LC50 of commercial formulations of abamectin (applied in a range from 6.8 to 0.84 mg L-1), Bacillus thuringiensis (640-5.1 mg L-1), fipronil (l,000-1 mg L-1), methoxyfenozide (9,600-60 mg L-1), spinosad (106-0.11 mg L-1), and triflumuron (24,000-2.4 mg L-1). Each larva was fed 1,5 µL of insecticide solution (30% honey solution mixed with the commercial formulations), and a control was fed with only 30% honey solution). The symptoms of the neurotoxic insecticides previous to death were body paralysis and cuticle darkening within the first 72 h; triflumuron was effective, but because of its form of action its symptoms took longer to appear (incomplete ecdysis, and/or partial rupture of the cuticle). The LC50 were determined by Probit analyses, were: spinosad, 0.29 mg L-1 abamectin, 1.40 mg L-1; fipronil, 3.34 mg L-1 and triflumuron, 11.83 mg L-1. Bacillus thuringiensis had no lethal effect on larvae in the range of concentrations evaluated. With methoxyfenozide, mortality was obtained only when using the highest concentration (9,600 mg L-1). [ABSTRACT FROM AUTHOR]

The ecdysone receptor (EcR) has been used to develop gene switches for conditional regulation of transgene expression in plants and humans. All EcR-based gene switches developed to date for use in plants are monopartate and require micromolar concentrations of ligand for activation of the transgene; this has limited the use of these gene switches. We have developed a Choristoneura fumiferana ecdysone receptor (CfEcR)-based two-hybrid gene switch that works through the formation of a functional heterodimer between EcR and the retinoid X receptor (RXR) upon application of the chemical ligand methoxyfenozide. Methoxyfenozide is already registered for field use with an excellent safety profile, and it has potential as a gene switch ligand for applications in the field. The receptor constructs were prepared by fusing DEF domains (hinge region plus ligand-binding domain) of CfEcR to the GAL4 DNA-binding domain and EF domains (ligand-binding domain) of ultraspiracle from Choristoneura fumiferana (CfUSP) or RXR from Locusta migratoria (LmRXR), Mus musculus (MmRXR) or Homo sapiens (HsRXR) to the VP16 activation domain. These receptor constructs were tested for their ability to induce expression of the luciferase gene placed under the control of 5× GAL4 response elements and −46 35S minimal promoter in tobacco, corn and soybean protoplasts and in transgenic Arabidopsis and tobacco plants. By adopting the two-hybrid format, the sensitivity of the CfEcR gene switch has been improved from micromolar to nanomolar concentrations of methoxyfenozide. The sensitivity of the CfEcR + LmRXR two-hybrid switch was 25 to 625 times greater than the monopartate gene switch, depending on the plant species tested. [ABSTRACT FROM AUTHOR]

The effect of the non-steroidal ecdysone agonist methoxyfenozide was evaluated against different developmental stages of the grape berry moth, Lobesia botrana Dennis & Schiffermuller (Lep, Tortricidae). Methoxyfenozide administered orally reduced the fecundity and fertility of adults treated with 1, 5 and 10 mg litre-1; longevity was not affected. An LC50 value of 4.5 mg litre-1 was obtained when applied to eggs of less than 1 day old. Surface treatment was more effective than when applied by spraying. Administered into the diet, methoxyfenozide had a larvicidal effect; older larvae were more susceptible than younger larvae, with LC50 values of 0.1 mg litre-1 for L1, 0.04 for L3 and 0.02 for L5. Larvae treated with sub-lethal doses throughout their lives did not emerge as adults at the highest doses (0.08, 0.04, 0.02 and 0.01 mg litre-1), with 65% and 40% emergence occurring for the lowest (0.005 and 0.0025 mg litre-1). Mortality occurred only in the larval stage. [ABSTRACT FROM AUTHOR]

Sublethal effects of the insect growth regulator methoxyfenozide were examined in oriental fruit moth, Grapholita molesta (Busck), in laboratory and field studies. In laboratory studies, oriental fruit moth larvae reared on diet amended with 0.1 ppm methoxyfenozide developed at the same rate as larvae reared on untreated diet, and paired moths reared as larvae from the same treated or untreated diets exhibited similar fecundity and fertility. Population growth differences over multiple generations were used to examine sublethal effects of methoxyfenozide on population dynamics in the field. Multiple single-tree cages were placed over apple (Malus spp.) trees treated with two applications of methoxyfenozide (70 g [AI/ha) and nontreated trees. Cages were infested at a single time point with virgin male and female oriental fruit moth adults, and population growth was evaluated by egg counts, shoot infestation, fruit damage, and larval counts over a 12-wk period. Significantly fewer eggs, larvae, and damaged fruit were found on compared with nontreated trees in 2001, Observed population differences may have been a result of direct mortality to eggs and larvae of the first generation rather than sublethal effects. In 2002, no differences were observed between treatments, but a heavy rain event shortly after the early infestation impacted the experiment. A late moth release treatment was tested in 2002 to examine the effects of residual methoxyfenozide 55 d after initial application. Significantly fewer eggs were found in the methoxyfenozide treatment compared with the control, but no differences existed among treatments in shoot infestation, percentage of damaged fruit, or larval populations. It was concluded direct mortality of eggs and larvae exposed to methoxyfenozide rather than sublethal effects were most important in reduction of subsequent generations. [ABSTRACT FROM AUTHOR]

The insect growth regulators (IGRs) tebufenozide and methoxyfenozide and the broad-spectrum insecticides azinphosmethyl, carbaryl, and fenpropathrin were compared for their activity against adult, egg, and larval stages of the grape berry moth, Endopiza viteana (Clemens) (Lepidoptera: Tortricidae), under laboratory and vineyard conditions. Adult mortality was not affected by exposure to field-equivalent rates of tebufenozide or methoxyfenozide on grape clusters, whereas all the broad-spectrum compounds significantly reduced adult survival, compared with the untreated controls. Surviving adult moths laid significantly more eggs on berries treated with the IGRs than on berries treated with any of the broad-spectrum insecticides. Survival of these eggs through to late larval and pupal stages was significantly lower on methoxyfenozide-treated grapes than on untreated grapes, and no pupae were found when grapes were treated with azinphosmethyl or fenpropathrin. Neither of the growth regulator insecticides limited egg eclosion or larval development by E. viteana when insecticides were applied before egg laying, whereas broad-spectrum insecticides were effective against both eggs and neonates at this timing. When applied after egg eclosion, all insecticide treatments significantly reduced survival of grape berry moth larvae. Under vineyard conditions, berries with 1-d-old residues of tebufenozide or methoxyfenozide received more E. viteana eggs than berries treated with broad-spectrum compounds. After aging for 7 or 14 d, no significant effects on E. viteana survival were detected among treatments. Whereas broad-spectrum insecticides provide control of multiple life stages of E. viteana, integration of tebufenozide or methoxyfenozide into vineyard management programs for control of this pest will be most successful if applications are timed for egg hatch. [ABSTRACT FROM AUTHOR]

The toxicity of two biorational insecticides, spinosad (Tracer) and methoxyfenozide (RH-2485), was tested against eggs, larvae, and pupae of the noctuid Spocloptera littoralis (Boisduval). In the first experiment, filter paper circles containing egg masses of two different age classes, young (<24 h old) and old (24-48 h old), were dipped in different concentrations of each insecticide diluted in either water or acetone. No ovicidal activity was recorded when insecticides were diluted in water. In contrast, when insecticides were diluted in acetone, both egg age classes generally showed a concentration-dependent response for both compounds. Mortality of larvae that hatched from both egg age classes was significantly increased, compared with control larvae, at all concentrations of both insecticides when diluted in water or acetone alike. The prevalence of mortality was similar with each insecticide. In the second experiment, third instars of S. littoralis were fed semisynthetic diet containing different concentrations of both insecticides. According to LC50 values, no significant differences were observed between spinosad (2.11 mg [AI]/kg diet) and methoxyfenozide (3.98 mg [AI]/kg diet) after 48 h of treatment, based on the overlap of 95% CL. Toxic effects on the mortality of pupae, adult emergence, and the prevalence of deformed adults after topical application on young pupae also were examined. Only methoxyfenozide caused pupal mortality and deformed adults. Our results suggest that spinosad and methoxyfenozide are potentially potent compounds for control of S. littoralis. [ABSTRACT FROM AUTHOR]

Hyposoter didymator (Thunberg) (Hymenoptera: Ichneumonidae) is an important larval parasitoid of several lepidopteran pests. Under laboratory conditions, the topical toxicity of selected insect growth regulator insecticides, azadirachtin (AZA), diflubenzuron (DFB), methoxyfenozide (MET), pyriproxyfen (PYR), and tebufenozide (TEB), and the naturalyte spinosad (SPIN) was tested against last (third)-instar larvae of this parasitoid up to the maximum field recommended concentration (MFRC). At MFRC, no abnormalities or mortality were scored for MET and TEB (IOBC toxicity classes 1). In contrast, AZA was slightly, PYR moderately, and DFB and SPIN harmful to the parasitoid (IOBC toxicity classes 2, 3, and 4, respectively). These ratings are the result of a reduction in rate of: (i) pupae formation, (ii) pupal mortality, (iii) adult longevity, (iv) parasitism, and (v) adult emergence. In addition, PYR and SPIN caused a direct mortality in the treated larvae. At sublethal concentrations, every insecticide but MET and TEB also affected these life history parameters of the parasitoid. The second part of the study focused on pharmacokinetic parameters important for the toxicological properties of these insecticides. We determined the recovery pattern of 14C-labeled DFB, MET, PYR, and TEB from parasitoid body, larval exuvia, and silken pupal cocoon after topical application to third-instar larvae. For MET, TEF, and PYR, >60% was recovered from the parasitoid body after 24 h, whereas this was only 12% for DFB, which was mostly detected in the silken cocoon (>60%). More than 25% PYR was also detected in the larval exuvia after three days. Data suggest that the silken pupal cocoon and the larval exuvia might play a role in eliminating insecticides. In conclusion, the current results indicated that MET and TEB are harmless to third larvae of H. didymator. In contrast, AZA, DFB, PYR, and SPIN are harmful and their use in IPM should be considered with caution until they are evaluated under field conditions. [Copyright &y& Elsevier]

Bioassays of Lacanobia subjuncta (Grote and Robinson) larvae established baseline LC50 values and identified the potential of reduced-risk, organophosphate replacement and naturally derived insecticides (eg chloronicotinyls, spinosyns, oxadiazines, insect growth regulators, microbial insecticides and particle films) to control this pest. The toxicities of these products were compared with those of organophosphate, carbamate, chlorinated cyclodiene and synthetic pyrethroid insecticides used in the management of lepidopteran pests in Washington apple orchards. Field trials were conducted comparing candidate insecticides to conventional alternatives. Several new insecticides (eg spinosad, methoxyfenozide, indoxacarb and an aluminosilicate particle film) proved to be effective for the management of L subjuncta. We summarize the goals and challenges of developing an integrated pest management program for new and resurgent pests as insecticide tools continue to change, and propose a hypothesis for the sudden increase in pest status of L subjuncta based on organophosphate tolerances. The role of novel insecticides with unique modes of action in resistance management and the encouragement of biological control are also discussed. [ABSTRACT FROM AUTHOR]

The phenology of oriental fruit moth, Grapholita molesta (Busck), on apple (Malus spp.) in North Carolina was studied using pheromone traps and egg sampling in abandoned and commercial orchards in 2000 and 2001, with subsequent development of an oviposition degree-day model and management studies in relation to codling moth, Cydia pomonella (L.), phenology. Oriental fruit moth eggs were found in greater numbers on leaves early and on fruit later in the growing season, on the top versus the bottom of the leaf surface, and on the calyx area versus the side or stem end of the fruit. A degree-day (DD) model to predict oriental fruit moth oviposition was developed based on temperature accumulations from peak moth trap capture of the first (overwintering) generation, by using 7.2 and 32.2°C as the temperature limits. The model predicted four ovipositing generations of oriental fruit moth with the second beginning 507 DD after peak moth catch. Using predictions of the oriental fruit moth and codling moth degree-day oviposition models, an experiment was conducted to determine the level of second generation oriental fruit moth control with methoxyfenozide applied under different scenarios for first generation codling moth. Methoxyfenozide was equally effective in managing codling moth and oriental fruit moth for all treatment timings. [ABSTRACT FROM AUTHOR]

Tebufenozide and methoxyfenozide have been previously shown to significantly reduce fecundity and cause vitellogenin accumulation in hemolymph of the codling moth Cydia pomonella L. In the present study, the effects of these ecdysone agonists, tebufenozide and methoxyfenozide, on the expression of ecdysone receptor (EcR), ultraspiracle protein (USP) and other proteins in the ovaries of C. pomonella L., were investigated at both the translational and/or transcriptional levels with an aim to elucidate the mechanisms by which the fecundity was reduced. Western and Northern blot analyses revealed that the expression of a 65 kDa (p65) EcR, and 60 and 64 kDa (p60 and p64) USP proteins were enhanced by tebufenozide and methoxyfenozide at both transcriptional and translational levels. Northern blot analysis indicated that the p65 EcR protein is encoded by EcRB1 transcript and that the p60 and p64 USP bands were the products of USP-1 transcript. Immunoprecipitation assays demonstrated that both the p60 and p64 USP coprecipitated with the p65 EcR and that p64 was a dominant USP to form complex with EcR. In addition, several other specific proteins were also identified and their expressions affected by the agonists. The data suggest that the ecdysone agonists regulate, via the EcR/USP complex, the expression of these specific proteins that might eventually lead to the inhibition of fecundity in the codling moth. [Copyright &y& Elsevier]

Abstract The attractiveness and responsiveness of the adult redbanded leafroller, Argyrotaenia velutinana (Walker), and obliquebanded leafroller, Choristoneura rosaceana (Harris), exposed to surfaces treated with the ecdysteroid agonist methoxyfenozide were investigated in wind tunnel and orientation tube assays. For both species, when males were exposed to water-treated surfaces for 48 h, and regardless of what treatment surfaces the females had been exposed to, the mean percentage recaptures of such treated males in the wind tunnel assays were always significantly greater than the recaptures of males that had been exposed to methoxyfenozide. However, exposure to surfaces treated with surfactant (Latron B-1956) also negatively affected the responsiveness (or attractiveness) of both moth species. In the orientation tube assays, males exposed to methoxyfenozide-treated surfaces always had significantly lower mean levels of individuals exhibiting sexual excitability, and the mean distances traveled upwind, regardless of females’ exposure. In addition, exposure to surfactant-treated surfaces did not affect mean levels of sexual excitibility and distances traveled upwind. The two bioassays demonstrated that male A. velutinana and C. rosaceana exposed to methoxyfenozide-treated surfaces were not as responsive to calling females (treated and non-treated) as were the non-treated males. Conversely, females exposed to methoxyfenozide-treated surfaces were just as attractive to non-treated males as water-treated females. It appears that a male's ability to respond to a calling female is more negatively affected by the ecdysone agonist than a female's ability to call and attract males. [ABSTRACT FROM AUTHOR]

We have developed an inducible gene expression system with potential for field application using the ecdysone receptor (EcR) from the spruce budworm and the non-steroidal EcR agonist, methoxyfenozide. Chimeric transcription activators were constructed with EcR ligand binding domain, GAL4 and LexA DNA binding domains, and VP16 activation domain. In the presence of methoxyfenozide, the transcription activators induced expression of the luciferase reporter gene cloned downstream of a promoter containing GAL4- or LexA-response element and a minimal 35S promoter. Low basal and high induced luciferase expression was optimized by cloning the activator and the reporter genes in different tandem orientations. Many transgenic Arabidopsis and tobacco plants were obtained with little or no basal expression in the absence of methoxyfenozide and inducible expression that was several fold higher than that observed with the constitutive 35S promoter. Moreover, gene expression was controlled over a wide range of methoxyfenozide concentration. Our results demonstrate that the inducible gene expression system based on the spruce budworm EcR ligand binding domain with methoxyfenozide as a ligand is very effective in regulating transgenes in plants. It is suitable for field applications because methoxyfenozide is commercially available and has an exceptional health and environmental safety profile. [ABSTRACT FROM AUTHOR]

A mutation in the maize Ms45 gene results in abortion of microspore development and a male-sterile phenotype. MS45 protein has been localized to the tapetum and maximally expressed in anthers at the early vacuolate stage of microspore development. Molecular complementation analysis determined that a transformed copy of the gene fully restored fertility to ms45 maize. In this report, using phenotypic complementation as an assay, chimeric transcriptional activators were expressed to regulate a gal:MS45 gene and test the ability of a multi-component system to restore male fertility. A high frequency of phenotypic complementation was observed when either C1-GAL4 or VP16-GAL4 activators were transcribed by promoters that expressed at a stage of anther development that precedes the early vacuolate stage of microsporogenesis. For the conditional regulation of male fertility, these transcriptional activators were modified by the addition of regions that include the ligand-binding domain from the European corn borer ecdysone receptor to generate the nuclear receptors C1-GAL4-EcR (CGEcR) and VP16-GAL4-EcR (VGEcR). These chimeric receptors were introduced with the gal:MS45 gene into ms45 maize, and in the absence of ligand, these plants were male sterile. In contrast, application of the ecdysone agonist, methoxyfenozide, to plants containing either a constitutive (Ubiquitin1) or anther-specific (maize 5126) VGEcR resulted in the restoration of fertility to ms45 plants grown in either the greenhouse or in the field. [ABSTRACT FROM AUTHOR]

A cell line derived from embryonic tissues of the European corn borer, Ostrinia nubilalis (UMC-OnE), was established in EX-CELL 401 medium containing 10% fetal bovine serum. The cells grew in suspension, and were mainly spherical in shape. The cell doubling times at the 17th and 79th passages were 56 and 36 h, respectively. DNA amplification fingerprinting showed that the DNA profile of the OnE cell line was different from that of the southwestern corn borer, Diatraea grandiosella (UMC-DgE), and that of the cotton bollworm, Helicoverpa zea (BCIRL-HZ-AM1). The OnE cell line was responsive to treatments of 20-hydroxyecdysone and the ecdysone agonists, methoxyfenozide (RH-2485) and tebufenozide (RH-5992). These compounds caused similar effects on the cells, which included cell clumping and decreased cell proliferation. The clumps were observed on the third day of incubation, and became larger after 7 d of incubation. After 168 h of incubation, methoxyfenozide and tebufenozide were 35 and 11 times more effective, respectively, in inhibiting proliferation of the OnE cells than was 20-hydroxyecdysone. [ABSTRACT FROM AUTHOR]

Spodoptera exigua is a worldwide pest afflicting edible vegetables and has developed varying levels of resistance to insecticides. Methoxyfenozide (MET), an ecdysteroid agonist, is effective against lepidopteran pests such as S. exigua. However, the mechanism of MET to S. exigua remains unclear. In this study, we analyzed the expression patterns of genes related to the ecdysone signaling pathway in transcriptome data treated with sublethal doses of MET and analyzed how expression levels of key genes affect the toxicity of MET on S. exigua. Our results demonstrated that 2639 genes were up-regulated and 2512 genes were down-regulated in S. exigua treated with LC30 of MET. Of these, 15 genes were involved in the ecdysone signaling pathway. qPCR results demonstrated that ecdysone receptor A (EcRA) expression levels significantly increased in S. exigua when treated with different doses of MET, and that the RNAi-mediated silencing of EcRA significantly increased mortality to 55.43% at 72 h when L3 S. exigua larvae were exposed to MET at the LC30 dose. Additionally, knocking down EcRA suppressed the most genes expressed in the ecdysone signaling pathway. The combination of MET and dsEcRA affected the expression of E74 and enhanced the expression of TREA. These results demonstrate that the adverse effects of sublethal MET disturb the ecdysone signaling pathway in S. exigua, and EcRA is closely related to MET toxic effect. This study increases our collective understanding of the mechanisms of MET in insect pests. [ABSTRACT FROM AUTHOR]

Methoxyfenozide is widely employed in paddy land and can flow out into the aquatic environment. The present study combines two approaches, namely, an aquatic microcosm simulation experiment and a multimedia fugacity model, to study the fate and distribution of methoxyfenozide in an aquatic microcosm containing water, sediment, aquatic plants, and zebrafish. The model results indicated that the simulated concentrations agreed with the observed values within one order of magnitude. The degradation rate was less than 7.0% in the three types of aquatic microcosms at 740 h in the model. Methoxyfenozide exhibited very high persistence in the aquatic microcosm. Water played a key role in the fate of methoxyfenozide, acting as a sink in the simulated aquatic environment, followed by sediment. Only approximately 2% of methoxyfenozide entered the organisms (zebrafish and Egeria densa Planch). Methoxyfenozide underwent a significant transport process between the water and sediment. The applications of multimedia fugacity models are useful for understanding the behaviors, fate, and transport of pesticides after their release into the environment and to facilitate risk assessment and management activities. Unlabelled Image • The microcosm combined with the fugacity model was first used to study the fate and distribution of methoxyfenozide. • The values obtained from the model calculations were in good agreement with values from the experiments. • Methoxyfenozide exhibits high persistence. Water served as a key sink for methoxyfenozide, followed by sediment. • This research will provide technical support for risk assessment and management activities of methoxyfenozide. [ABSTRACT FROM AUTHOR]

The widespread use of pesticides is one of the major health and environmental concerns. There is an urgent need to develop improved analytical methodologies as well as for a deep insight into the mechanism behind the interaction between pesticides and the nucleic acids of living organisms. In the present work, the electrochemical behavior of the diacylhydrazine insecticide methoxyfenozide (Met) on a boron-doped diamond electrode and its interaction with double-stranded DNA (dsDNA) were investigated for the first time by applying cyclic voltammetry and square-wave (SW) voltammetry. The influence of numerous factors, such as composition of the supporting electrolyte, its pH, and voltammetric parameters, was analyzed. Under the optimized conditions, a simple and sensitive SW voltammetric procedure was developed for the determination of Met. Quantification was found to be linear from 5.0 × 10−7 to 7.0 × 10−5 mol L−1, with a detection limit of 1.4 × 10−7 mol L−1. The application of the developed voltammetric methodology for the analysis of tap water, river water and grape juice was tested with spiked samples. Studies on the interaction between methoxyfenozide and double-stranded salmon sperm DNA were conducted using both voltammetric and spectroscopic techniques. The results obtained indicated the dominance of the intercalative binding mode between Met and dsDNA. The binding constants of the formed complex were calculated based on the changes in the electrochemical and spectroscopic behaviors of Met in the presence of dsDNA. • First electroanalytical study of insecticide methoxyfenozide. • Successful application of proposed methodology in spiked samples analysis • Cheap, fast, and environmentally friendly methodology. • Electrochemical and spectroscopic assay of the interaction between Met and dsDNA. • The dominance of the intercalative binding mode between Met and dsDNA. [ABSTRACT FROM AUTHOR]

Methoxyfenozide and metaflumizone are insecticides used on Chinese broccoli to prevent insects and increase yield. However, the residues are potentially harmful to the environment and consumers. In this study, the quick, easy, cheap, effective, rugged, safe method with high-performance liquid chromatography with tandem mass spectrometry was modified and validated for determination of methoxyfenozide and metaflumizone in Chinese broccoli. The clean-up efficiency of different sorbents including C 18 , primary secondary amine, graphitized carbon black, and carbon nanofiber was compared. Recoveries of the validated method were 71.8-94.6% with relative standard deviations of 1.5-3.2% and the limits of quantification were 0.01 and 0.005 mg/kg for methoxyfenozide and metaflumizone, respectively. A storage stability test showed almost no degradation of methoxyfenozide in Chinese broccoli, however, the degradation rate of metaflumizone was 22.9% after 10-wk storage at -20°C. In field trials in four producing regions, the dissipation of both methoxyfenozide and metaflumizone in Chinese broccoli was fast, with half-lives of only 1.0-5.1 and 0.7-2.5 days, respectively. Terminal residues after application of the two pesticides were all below 1.0 mg/kg after 5 days., (© 2021 Wiley-VCH GmbH.)

Honey bees (Apis mellifera) are highly valued pollinators that help to ensure national food security in the United States, but reports of heavy annual losses to managed colonies have caused concerns and prompted investigations into the causes of colony losses. One factor that can negatively affect honey bee health and survival is agrochemical exposure. Investigations into the sublethal effects of agrochemicals on important metrics of colony health such as reproduction and queen fecundity has been limited by the availability of targeted methods to study honey bee queens. This work investigates the effects of three insect growth regulators (IGR), a class of agrochemicals known to target pathways involved in insect reproduction, on honey bee queen oviposition, egg hatching, and worker hypopharyngeal development in order to quantify their effects on the fecundity of mated queens. The reported results demonstrate that none of the IGRs affected oviposition, but all three affected egg eclosion. Worker bees consuming methoxyfenozide had significantly larger hypopharyngeal glands at two weeks of age than bees not fed this compound. The results suggest that although IGRs may not exhibit direct toxic effects on adult honey bees, they can affect larval eclosion from eggs and the physiology of workers, which may contribute to colony population declines over time. Image 1 • Insect growth regulators can negatively affect honey bee embryo survival without impacting egg production rates. • Methoxyfenozide affects honey bee worker hypopharyngeal development. • Although IGRs may not cause mortality, they can affect adult honey bee physiology and fecundity in ways that may be detrimental to colony function and longevity. [ABSTRACT FROM AUTHOR]

Indoxacarb is an important insecticide for the selective control of Helicoverpa armigera. It can be bioactivated to the more effective N-decarbomethoxylated indoxacarb (DCJW) by esterases in pests. It was observed that both field and laboratory selected populations of H. armigera showed negative cross-resistance between indoxacarb and methoxyfenozide. The Handan population exhibited moderate resistance to indoxacarb, but was susceptible to methoxyfenozide; the Baoding and Yishui populations exhibited moderate resistance to methoxyfenozide, but they were susceptible to indoxacarb. Moreover, the toxicity of indoxacarb was enhanced 1.83-fold in the laboratory methoxyfenozide-resistant H. armigera, and susceptibility to methoxyfenozide was increased 2.81-fold in the laboratory indoxacarb-resistant H. armigera. In vivo, DCJW concentrations in the susceptible and methoxyfenozide-selected (laboratory methoxyfenozide-resistant) populations were 4.59- and 4.31-fold greater than in the indoxacarb-resistant Handan population 1 h after dosing. After 2 h, the highest concentrations of DCJW and indoxacarb appeared in the methoxyfenozide-selected population. Meanwhile, increased carboxyl esterase (CarE) and decreased glutathione S-transferase (GST) activities were observed in the methoxyfenozide-selected population. However, the indoxacarb-selected (laboratory indoxacarb-resistant) and Handan populations showed a higher disappearance of indoxacarb and DCJW, and the activity of cytochrome P450 mono-oxygenase in these populations were significantly increased. This study showed that the improved toxicity of indoxacarb, as observed in the methoxyfenozide-selected H. armigera, was correlated with increased CarE activity, decreased GST activity, and the in vivo accumulation of indoxacarb and DCJW. The significantly increased cytochrome P450 activity and higher disappearance of indoxacarb and DCJW in indoxacarb-resistant H. armigera resulted in the decreased toxicity of indoxacarb. [ABSTRACT FROM AUTHOR]

Insecticide based vector control tools such as insecticide treated bednets and indoor residual spraying represent the cornerstones of malaria control programs. Resistance to chemistries used in these programs is now widespread and represents a significant threat to the gains seen in reducing malaria-related morbidity and mortality. Recently, disruption of the 20-hydroxyecdysone steroid hormone pathway was shown to reduce Plasmodium development and significantly reduce both longevity and egg production in a laboratory susceptible Anopheles gambiae population. Here, we demonstrate that disruption of this pathway by application of the dibenzoylhydrazine, methoxyfenozide (DBH-M), to insecticide resistant An. coluzzii , An. gambiae sl and An. funestus populations significantly reduces egg production in both topical and tarsal application. Moreover, DBH-M reduces adult longevity when applied topically, and tarsally after blood feeding. As the cytochrome p450s elevated in pyrethroid resistant Anopheles only bind DBH-M very weakly, this compound is unlikely to be subject to cross-resistance in a field-based setting. Manipulation of this hormonal signalling pathway therefore represents a potential complementary approach to current malaria control strategies, particularly in areas where high levels of insecticide resistance are compromising existing tools. Image 1 • Topical application of methoxyfenozide is effective at sterilising insecticide resistant populations of African Anophelines. • Topical application also significantly decreases lifespan. • Tarsal application with the adjuvant RME similarly sterilises and shortens life span post-blood feeding. • Cytochrome p450s, major insecticide metabolisers, don't show affinity for methoxyfenozide indicating no cross-resistance. • Methoxyfenozide has potential importance in a field-setting as a complementary vector control tool. [ABSTRACT FROM AUTHOR]

Abstract: The insecticidal activity of two ecdysone agonists, methoxyfenozide (RH-2485) and halofenozide (RH-0345), was tested against last-instar larvae of the natural predator Harmonia axyridis (Col., Coccinellidae). In addition, the relative weight gain of the larvae after application was followed. Both products proved to be equally toxic at concentrations of 25, 50 and 100 mg/l. The ecdysteroidal activity of the compounds caused premature induction of larval moulting, cessation of feeding and incomplete pupation in affected larvae. Although, compared with previous results with methoxyfenozide and halofenozide in target pests such as the Colorado potato beetle, Leptinotarsa decemlineata, these compounds caused mortality only after application at relatively high concentrations. [ABSTRACT FROM AUTHOR]

Background: In holometabolous insects, the major developmental transitions - larval molting and pupation - are triggered by a pulse of 20-hydroxyecdysone (20E) and coordinated by juvenile hormone. Methoxyfenozide (MF), an ecdysteroid agonist, represents a new class of insect growth regulators and is effective against lepidopteran pests. Fushi-tarazu factor 1 (FTZ-F1) is an ecdysone-inducible transcription factor. To date, the effect of MF on 20E-response genes remains unclear, and we speculate the involvement of FTZ-F1 in MF's growth regulating effect., Results: MF at LC 25 and LC 10 caused severe ecdysis failure in Helicoverpa armigera, extended their larval duration, lowered their pupal weight, and reduced the respiratory, pupation and emergence rates. Furthermore, sublethal doses of MF inhibited ecdysteroidogenesis and lowered the intrinsic 20E titer, but showed an inductive effect on 20E-response genes including HaFTZ-F1. HaFTZ-F1, predominantly expressed in larval epidermis, was markedly upregulated before or right after larval ecdysis, and maintained a high level in prepupal stage. Knockdown of HaFTZ-F1 in 4th-instar larvae severely impaired larval ecdysis, whereas its knockdown in final-instar larvae caused abnormal pupation. Moreover, knocking down HaFTZ-F1 downregulated three critical ecdysteroidogenesis genes, lowered 20E titer, and suppressed the expression of 20E receptors and 20E-response genes. The introduction of 20E into HaFTZ-F1-RNAi larvae partly relieved the negative effects on the 20E-induced signaling cascade., Conclusion: Our findings reveal the adverse effects of sublethal doses of MF on the development of H. armigera and elucidate the resulting perturbations on the 20E-induced signaling cascade; we propose that HaFTZ-F1 regulates ecdysis and pupation by mediating 20E titer and its signaling pathway. © 2020 Society of Chemical Industry., (© 2020 Society of Chemical Industry.)

In compliance with Article 43 of Regulation (EC) No 396/2005, the EFSA received from the European Commission a mandate to provide its reasoned opinion on the existing maximum residue levels (MRLs) for methoxyfenozide which might lead to consumers intake concerns on the basis of the new toxicological reference values agreed upon by Member States (MSs) on 13 December 2018. In order to identify the MRLs of potential concern that require a more detailed assessment, EFSA screened the existing MRLs for methoxyfenozide, considering the new toxicological reference values and an acute risk could not be excluded for eight commodities. A fall-back MRL was proposed for tomatoes and the MRL for citrus fruit could be confirmed considering the use of a peeling factor. No other fall-back good agricultural practices (GAPs) were received, and thus, a lowering of the MRLs for peaches, apples, pears and broccoli is proposed., (© 2020 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.)

Background: Methoxyfenozide possesses efficacy against a variety of lepidopteron pests, including the major pests in cauliflower and tea, so it is of great importance to generalize the practical use of methoxyfenozide in the field., Results: An efficient method was developed and validated in both vegetable matrix and extract-rich matrix (cauliflower and tea) using modified QuEChERS combined with UPLC/MS/MS analysis. The recoveries in cauliflower, made tea and tea shoots ranged from 94.5 to 108.0%, from 85.0 to 91.6% and from 77.3 to 82.0% respectively, with relative standard deviations (RSDs) below 17.3% in all cases. The field results showed that methoxyfenozide dissipated in cauliflower with half-life (t 1/2 ) at 2.5-3.5 days and in tea with t 1/2 at 1.2 days. Combining the above experimental data and statistical food intake values, the risk quotient (RQ) values were significantly lower than 1., Conclusion: The quantification method of methoxyfenozide in cauliflower or tea has not been established until this study. The dissipation and dietary exposure risk assessment of methoxyfenozide in cauliflower and tea were investigated in the field. Methoxyfenozide dissipated rapidly in cauliflower despite different climates, and it dissipated faster in tea. The dietary risk of methoxyfenozide through cauliflower or tea was negligible to humans. This study not only provides guidance for the safe use of methoxyfenozide but also serves as a reference for the establishment of maximum residue limits (MRLs) in China. © 2019 Society of Chemical Industry., (© 2019 Society of Chemical Industry.)