Chitin Synthesis Inhibitor Diflubenzuron Research Articles

Lethal effects of insecticides on adults of Eriopis connexa (Germar, 1824) (Coleoptera: Coccinellidae)

The predator Eriopis connexa (Germar, 1824) (Coleoptera: Coccinelidae), is an important biological control agent in wheat fields. In this work, we evaluated the lethal effects of fifteen insecticides used in the wheat crop on adults of the predator E. connexa, using the methodology proposed by International Organization for Biological and Integrated Control (IOBC). Sublethal effects have not been evaluated. The insecticides were sprayed on glass plates that were used to set up the exposure arenas. The chitin synthesis inhibitors diflubenzuron SC, diflubenzuron WP, lufenuron, triflumuron, and the pyrethroid beta-cyfluthrin were considered harmless to adults of the predator and should have their use prioritized. On the other hand, methomyl, lambda-cyhalothrin, thiamethoxam + lambda-cyhalothrin A and B were considered harmful to E. connexa and should be avoided when adults of the predator are present in the crop.

Statewide Efficacy Assessment of Insect Growth Regulators Against Aedes albopictus (Diptera: Culicidae) in Sabah, Malaysia: An Alternative Control Strategy?

The efficacy of three groups of insect growth regulators, namely juvenile hormone mimics (methoprene and pyriproxyfen), chitin synthesis inhibitors (diflubenzuron and novaluron), and molting disruptor (cyromazine) was evaluated for the first time, against Aedes albopictus Skuse (Diptera: Culicidae) larvae from 14 districts in Sabah, Malaysia. The results showed that all field populations of Ae. albopictus were susceptible towards methoprene, pyriproxyfen, diflubenzuron, novaluron, and cyromazine, with resistance ratio values ranging from 0.50-0.90, 0.60-1.00, 0.67-1.17, 0.71-1.29, and 0.74-1.07, respectively. Overall, the efficacy assessment of insect growth regulators in this study showed promising outcomes and they could be further explored as an alternative to conventional insecticides.

Exposing Northern shrimp (Pandalus borealis) to fish feed containing the antiparasitic drug diflubenzuron caused high mortality during molting

ABSTRACTUse of the chitin synthesis inhibitor diflubenzuron (DFB) as an antiparasitic drug in salmon aquaculture raises concern over its impact on marine ecosystems. Further, global drivers, such as ocean warming and acidification (OAW), may increase the toxicity of hazardous substances including DFB. The aim of the present study was to examine the combined effects of DFB-medicated salmon feed on ovigerous Northern shrimp (Pandalus borealis) under Control (pHNBS 8.0, 7.0ºC) and OAW conditions (pHNBS 7.6, 9.5ºC). DFB-exposed shrimp consumed on average 0.1–0.3 g medicated feed during the 2-week exposure period, and high mortality (61–73%) was documented at both environmental conditions. There was no significant interaction between OAW and DFB. Only 2–7% of DFB-exposed shrimp molted successfully compared to 65% in Control and 63% in OAW. The shrimp molted earlier (shorter intermolt period) and exhibited higher feeding rate at OAW compared to Control conditions. An additional experiment, where female shrimp were exposed to DFB closer to molting, noted increased mortality after only 4 d exposure, and successful molting for some shrimp after 2 to 3 weeks of depuration. High mortality of shrimp exposed to DFB-medicated feed indicates that the use of this feed in aquaculture could affect local shrimp populations.

Insecticides based on insect chitin synthesis inhibitors

The following benzoylurea chitin synthesis inhibitors—diflubenzuron used in Dimilin and Gerold formulations, lufenuron registered as Match, and a Lufox combined insecticide based on two active ingredients, lufenuron and fenoxycarb (juvenoid)—have been tested. The effectiveness of the Dimilin insecticide in regulating the codling moth (Cydia pomonella L.) population was assessed in Tambov oblast, Rostov-on-Don oblast, and Krasnodar krai in 2010–2011. Reduction of damage in the harvested fruit at the maximum application rate comprised 90–100% in all the regions. The biological effectiveness of the Gerold preparation in regulating the complex population consisted of leafrollers and currant bud moths (Incurvaria capitella Cl.) and comprised 85–87% in the Tambov oblast and reached 100% in the Orel and Leningradskaya oblasts (2012–2014). The field tests to assess the biological effectiveness of the Match insecticide in regulating the population of cotton worm (Heliothis armigera Hb) in Krasnodar krai and Astrakhan oblast proved that the preparation was capable to reduce the number of the cotton worm larvae on tomato plants (up to 99.0%). The Lufox combined insecticide effectively reduced the larval population of codling moth and, hence, the damage of apple fruit in Krasnodar krai and Rostov-on-Don oblast (up to 96.8%). The tested plant protection products are recommended for use in the ecologized systems.

Identification, characterization and functional analysis of a chitin synthase gene in the brown citrus aphid, Toxoptera citricida (Hemiptera, Aphididae).

Chitin synthase (CHS) is a crucial enzyme involved in the final step of the insect chitin biosynthetic pathway. In this study, we cloned the full-length cDNA sequence of a chitin synthase gene (TCiCHS) from the brown citrus aphid, Toxoptera citricida, an important citrus pest and the main vector of citrus tristeza virus worldwide. TCiCHS was expressed during the entire lifecycle and in all insect tissues examined. Expression was highest in first-second-instar nymphs, nymph-adult transitions and in the abdomen (6.7-fold higher than head). Embryos had a higher expression level than the integument. Fourth-instar nymphs were exposed to 5 and 500 mg/l concentrations of the chitin synthesis inhibitor diflubenzuron (DFB) for 48 h and had the highest mortality at the 500 mg/l concentration. The mRNA expression levels of TCiCHS were significantly enhanced upon the exposure of nymphs to both low and high DFB concentrations. Silencing of TCiCHS occurred through plant-mediated double-stranded RNA (dsRNA) feeding. Most dsRNA-fed nymphs were unable to moult to the next stage, and the expression of TCiCHS decreased 48% compared with controls. These results demonstrate that TCiCHS plays an important role in nymph to adult development, is possibly help identify molecular targets for To. citricida control.

Chitin is a necessary component to maintain the barrier function of the peritrophic matrix in the insect midgut

In most insects, the peritrophic matrix (PM) partitions the midgut into different digestive compartments, and functions as a protective barrier against abrasive particles and microbial infections. In a previous study we demonstrated that certain PM proteins are essential in maintaining the PM's barrier function and establishing a gradient of PM permeability from the anterior to the posterior part of the midgut which facilitates digestion (Agrawal et al., 2014). In this study, we focused on the effects of a reduction in chitin content on PM permeability in larvae of the red flour beetle, Tribolium castaneum. Oral administration of the chitin synthesis inhibitor diflubenzuron (DFB) only partially reduced chitin content of the larval PM even at high concentrations. We observed no nutritional effects, as larval growth was unaffected and neutral lipids were not depleted from the fat body. However, the metamorphic molt was disrupted and the insects died at the pharate pupal stage, presumably due to DFB's effect on cuticle formation. RNAi to knock-down expression of the gene encoding chitin synthase 2 in T. castaneum (TcCHS-2) caused a complete loss of chitin in the PM. Larval growth was significantly reduced, and the fat body was depleted of neutral lipids. In situ PM permeability assays monitoring the distribution of FITC dextrans after DFB exposure or RNAi for TcCHS-2 revealed that PM permeability was increased in both cases. RNAi for TcCHS-2, however, led to a higher permeation of the PM by FITC dextrans than DFB treatment even at high doses. Similar effects were observed when the chitin content was reduced by feeding DFB to adult yellow fever mosquitos, Aedes aegypti. We demonstrate that the presence of chitin is necessary for maintaining the PM's barrier function in insects. It seems that the insecticidal effects of DFB are mediated by the disruption of cuticle synthesis during the metamorphic molt rather than by interfering with larval nutrition. However, as DFB clearly affects PM permeability, it may be suitable to increase the efficiency of pesticides targeting the midgut.

Toxicological assessment of the anti-salmon lice drug diflubenzuron on Atlantic cod Gadus morhua

Increasing use of the chitin synthesis inhibitor diflubenzuron against the ectoparasitic salmon louse Lepeophtheirus salmonis in marine aquaculture has raised concerns over its environmental impacts. This study evaluated how diflubenzuron affects Atlantic cod Gadus morhua, a fish species often found near Atlantic salmon Salmo salar farms, focusing on uptake kinetics and hepatic transcriptional responses. Two experiments were conducted, one time-series trial in which the fish were given a daily dose (3 mg kg-1 fish) of diflubenzuron for 14 d followed by a 3 wk depuration period, and one dose-response trial with increasing concentrations (3, 10 and 50 mg kg-1 fish). The highest diflubenzuron concentrations were found in the liver at Day 15. No detectable levels of diflubenzuron were found in liver or muscle 3 wk after the end of the treatment. At the molecular level, small effects of diflubenzuron treatment on gene transcription were observed. In the time-series experiment, the strongest effects were seen at Day 8, with 2 transcripts being upregulated (bclx2 and cpt1a) and 8 transcripts being downregulated (gstp1, gstm1, gstt1, ugt1a, nat2, cat, p53 and slc16a9a). Five transcripts (cyp3a, cpt1a, ptgs2, elovl5 and mapk1) responded significantly to diflubenzuron exposure in the dose-response experiment. This study shows that diflubenzuron can be taken up by Atlantic cod, that it is rapidly cleared from the body and that when present this pharmaceutical causes only small effects on the expression of genes involved in detoxification pathways. Taken together, our data suggest that accumulated diflubenzuron at the levels studied would have a relatively small effect on wild Atlantic cod.

Comparative Studies on Effects of Three Chitin Synthesis Inhibitors on Common Malaria Mosquito (Diptera: Culicidae)

Toxicities of three chitin synthesis inhibitors (diflubenzuron, nikkomycin Z and polyoxin D) were evaluated using second instars of the common malaria mosquito, Anopheles quadrimaculatus Say (Diptera: Culicidae). Neither nikkomycin Z nor polyoxin D at 50 microg/liter caused significant larval mortality, although they reduced the body weight of the survivors by 20.5 and 33.8%, respectively, in 48 h. In contrast, exposures of the larvae to diflubenzuron at 12.5 microg/liter for 48 h resulted in 86.7% larval mortality and reduced the body weight of the survivors by 29.1%. Exposure of the pupae (<12 h old) to diflubenzuron at 100 microg/liter for 48 h caused 18.9% pupal mortality and consequently reduced the adult emergence by 24.7% from the surviving pupae. Furthermore, exposure of third instars to diflubenzuron at 4, 20, 100, and 500 microg/liter for 24 h resulted in the reduction of larval chitin contents by 4.25, 33.2, 35.2, and 57.7%, respectively. Such an effect seemed to be associated with only cuticular chitin synthesis because the same exposures did not significantly affect chitin contents in the guts. Our results indicated that diflubenzuron was highly toxic to second instars by not only causing high larval mortality but also by affecting their growth. Diflubenzuron was also fairly toxic to pupae by not only causing pupal mortality but also affecting the adult emergence. Our results suggest that diflubenzuron might affect only chitin synthesis in the cuticle but not in the peritrophic matrix, which is probably due to diflubenzuron's direct contact to mosquito larvae in water, slow distribution in insect body, rapid degradation in the insect gut, or a combination.

Laboratory Evaluation of Diflubenzuron as a Feed-Through for Control of Immature Sand Flies (Diptera: Psychodidae)

The benzoylurea chitin synthesis inhibitor diflubenzuron was evaluated as a rodent feed-through for the control of immature stages of Phlebotomus papatasi Scopoli (Diptera: Psychodidae). The development and survival of second instars of P. papatasi larvae that were fed feces from Syrian hamsters, Mesocricetus auratus, that had been fed a diet containing 0, 8.97, 89.7, or 897 ppm diflubenzuron was evaluated. No pupation or adult emergence occurred when larvae were fed feces from hamsters that were fed diets containing diflubenzuron. The mortality of sand flies fed feces from treated hamsters was coincident with pupation of the controls, suggesting a specific effect on the larval-to-pupal molt. The results of this study suggest that a control strategy using rodent baits containing diflubenzuron for phlebotomine sand flies and zoonotic cutaneous leishmaniasis may be possible.

Laboratory Evaluation of Diflubenzuron as a Feed-Through for Control of Immature Sand Flies (Diptera: Psychodidae)

The benzoylurea chitin synthesis inhibitor diflubenzuron was evaluated as a rodent feed-through for the control of immature stages of Phlebotomus papatasi Scopoli (Diptera: Psychodidae). The development and survival of second instars of P. papatasi larvae that were fed feces from Syrian hamsters, Mesocricetus auratus, that had been fed a diet containing 0, 8.97, 89.7, or 897 ppm diflubenzuron was evaluated. No pupation or adult emergence occurred when larvae were fed feces from hamsters that were fed diets containing diflubenzuron. The mortality of sand flies fed feces from treated hamsters was coincident with pupation of the controls, suggesting a specific effect on the larval-to-pupal molt. The results of this study suggest that a control strategy using rodent baits containing diflubenzuron for phlebotomine sand flies and zoonotic cutaneous leishmaniasis may be possible.

Uptake and distribution of three insect growth regulators — diflubenzuron, flucycloxuron and halofenozide — in pupae and adults ofTenebrio molitor

In a topical bioassay with pupae and adults of the mealwormTenebrio molitor (Coleoptera: Tenebrionidae), an important pest in stored products worldwide, three insect growth inhibitors (IGRs) were tested: the two chitin synthesis inhibitors diflubenzuron and flucycloxuron, and the ecdysone agonist halofenozide (RH-0345). To address differences in toxicity, we determined with the use of a14C-labeled isotope, the pattern of absorption through the cuticle and then the distribution in the different body tissues of gut, hemolymph, integument and reproductive organs (ovaries, testis). The rate of absorption through the cuticle was highest for flucycloxuron, and this concurred with its high toxicity. In general, the patterns of absorption followed a similar trend in young and old pupae and in the adult stage. Accumulation in the reproductive system of females and males was relatively high, which may explain the strong reproductive effects of the IGRs tested. In addition, we recorded large differences in the clearance of insecticide from the insect bodyvia the excrements, with the chitin synthesis inhibitors exhibiting higher rates than halofenozide.

Disruption of Reproductive Activity of &lt;I&gt;Coptotermes formosanus&lt;/I&gt; (Isoptera: Rhinotermitidae) Primary Reproductives by Three Chitin Synthesis Inhibitors

Effects of the chitin synthesis inhibitors (CSIs) diflubenzuron, hexaflumuron, and lufenuron on the Formosan subterranean termite, Coptotermes formosanus Shiraki, primary reproductives were studied in the laboratory. Incipient colonies were established by collecting and pairing C. formosanus alates and placing them in dishes containing an artificial diet. Three groups of 56 colonies each were fed with a diet containing 10 ppm of one of each of the CSIs and were compared with a control group fed with an untreated diet. All eggs oviposited by treated young queens failed to hatch at the end of 6 mo. Estimated queen fecundity was significantly lower in the lufenuron treatment compared with the control group. Fecundity of hexaflumuron-treated queens did not differ significantly from that of queens from the control group and the other treatments. Adult mortality was significantly higher in the diflubenzuron and lufenuron treatments than in the control group at the end of 6 mo., but not in the hexaflumuron treatment. All the pairs died within 8 mo. in the diflubenzuron and lufenuron treatments, even after treatment was suspended at the end of 6 mo. Mortality in the hexaflumuron treatment was significantly higher than in the control group by the end of 9 mo. The three CSIs tested eliminated reproduction in C. formosanus by preventing egg hatching and induced adult mortality. Possible mechanisms by which CSIs induce termite adult death are discussed.

Efficacy of diflubenzuron plus methoprene against Sitophilus oryzae and Rhyzopertha dominica in stored sorghum

The efficacy of diflubenzuron (1 mg kg −1)+methoprene (1 mg kg −1) against Sitophilus oryzae (L.) and Rhyzopertha dominica (F.) in sorghum was evaluated in a silo-scale trial in southeast Queensland, Australia. Sorghum is normally protected from a wide range of insects by mixtures of grain protectants. The chitin synthesis inhibitor diflubenzuron was evaluated as a potential new protectant for S. oryzae in combination with the juvenile hormone analogue methoprene, which is already registered for control of R. dominica. Sorghum (ca 200 t) was treated after harvest in 2000 and assessed for treatment efficacy and residue decline during 6.5 months storage. The reproductive capacity of S. oryzae and R. dominica was greatly reduced in bioassays of treated sorghum throughout the trial, and efficacy remained relatively stable during the trial. An initial exposure of S. oryzae adults to treated sorghum for 2 weeks reduced F 1 progeny production of all strains by 80.8–98.8%, but a second exposure of 4 weeks reduced F 1 progeny production by 98.5–100%. In addition, the reproductive capacity of any S. oryzae progeny produced was greatly reduced. Exposure of R. dominica adults to treated sorghum for 2 weeks reduced F 1 progeny production of all strains by 99.6–100%, including a methoprene-resistant strain. The results indicate that S. oryzae or R. dominica adults invading sorghum treated with diflubenzuron (1 mg kg −1)+methoprene (1 mg kg −1) would be incapable of producing sustainable populations.

Effects of diflubenzuron on the integument of fifth instarGalleria mellonella larvae

Effects of the chitin synthesis inhibitor diflubenzuron (DFB) on the integument of 5th instarGalleria mellonella L. larvae were investigated. When larvae were fed with semi-artificial diets containing 250, 500 and 1000 ppm of the compound, DFB affected the integument. The affected larvae failed to ecdysis, their cuticle was ruptured, lost haemolymph and blackened. In treated larvae, cuticle deposition was disrupted and the cuticle thickness was decreased by ∼50% compared with the untreated control, particularly at day 3 1/2. However, statistically there was no significant difference among the three concentrations (P>0.05). This may indicate that all three concentrations are equally effective in decreasing level. DFB at 500 and 1000 ppm also affected the epidermal cells and caused the occurrence of vacuoles.

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

AbstractThe 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 Diflubenzuron and Teflubenzuron in Larvae of Beet Armyworm (Lepidoptera: Noctuidae)

Spodoptera exigua (Hubner), a serious lepidopterous pest in greenhouse crops in the Netherlands, is rather insensitive to the chitin synthesis inhibitor diflubenzuron. The LC 50 for fourth and fifth instars treated with diflubenzuron were 295 and 16 mg/liter, respectively ; for teflubenzuron, LC 50 's were 42 and 7 mg/liter, respectively. To optimize the use of benzoylphenyl urea in S. exigua larvae, a combination with four synergists was studied. The best results were obtained with profenofos and S,S,S-tributylphosphorotrithiote (DEF), both of which are hydrolase inhibitors. These synergists enhanced the toxicity of diflubenzuronin fourth instars nine and six fold, respectively. Diethylmaleate, a glutathion S-transferase inhibitor, decreased the LC 50 five fold, whereas piperonyl butoxide, an oxidase inhibitor, was responsible for only a three-fold reduction

Effects of the chitin synthesis inhibitor diflubenzuron on development of Ascaris suum and Haemonchus contortus

The potential of the chitin synthesis inhibitor diflubenzuron (DFB) to alter the development of the parasitic nematodes ( Ascaris suum and Haemonchus contortus was investigated. DFB given orally (10 mg kg −1 per day for 30 days) to sheep inoculated with H. contortus infective larvae did not prevent the establishment of adults or affect fecal egg output. However, there was a significant (>90%) decrease in the number of infective larvae recovered from fecal cultures derived from lambs harboring H. contortus adults that were treated with DFB. DFB did not affect egg hatching. Oral administration (10 mg kg −1 per day for 20 days) of DFB to swine harboring adult A. suum adults had no effect on the adult worm burden or on egg morphology, but eggs removed from worms obtained from DFB-treated swine contained less chitin than eggs removed from untreated control swine. DFB also inhibited chitin synthesis in vitro in the isolated reproductive tract of A. suum adults. These results indicate that DFB at high doses can inhibit the subsequent development of H. contortus larvae in the feces. Since H. contortus larvae lack chitin, DFB may act on these larvae by a mechanism independent of a direct effect on chitin synthesis.

Resistance to Diflubenzuron in the Codling Moth (Lepidoptera: Tortricidae)

Codling moth, Cyndia Pomona ella (L.), from pear in the Bedford, Oreg., area showed resistance to the chitin synthesis inhibitor diflubenzuron in laboratory and orchard studies. In the orchard this was shown in 1984 and 1985 as 54.5-63.6% damage at harvest compared with 0.8-2.0% damage occurring in another similarly treated orchard. Concentration-mortality studies later in 1985 and early 1986 showed about a 2-fold difference in susceptibility (LC30= 52.6 and 59.1 mg [All/liter versus 37.9 and 28.2 mg [All/liter) between the two populations based on the response of eggs of the F3. and F4. laboratory-reared generations to residues on the treated fruits. When results from exposure of eggs to a single discriminating concentration of diflubenzuron (112.3 mg [All/liter) were compared, the diflubenzuron-resistant population showed a significantly higher survival (≤29.2-fold). Studies in the same orchard in 1986 again showed a lack of control (35% damage) with a normal concentration. Higher concentrations of 2 times and 4 times normal showed 15% and 8% damage, respectively. This is the first report of resistance to diflubenzuron in the codling moth. This occurrence of resistance is unusual because it appeared the first time this population was exposed to diflubenzuron without previous exposure to related chemicals, whereas the population in another similarly treated orchard retained a high degree of susceptibility after approximately 8 yr of exposure to diflubenzuron. The causes or mechanisms for this develpment of resistance to diflubenzuron in the codling moth remain to be determined.

Membrane binding and uptake of diflubenzuron in a cell line from Manduca sexta (L.)

AbstractThe binding and accumulation of the chitin synthesis inhibitor diflubenzuron (DFB) by a cell line derived from embryonic tissue of the tobacco hornworm, Manduca sexta (L.), was analyzed. A rapid and reversible binding to viable and nonviable cells suspended in the culture medium was observed at soluble concentrations of DFB for short exposure periods. Scatchard analysis gave no indication of a saturable uptake mechanism. The DFB‐binding capacity of intact cells was found to be similar to that of a crude membrane preparation (70,000g pellet); however, plasma membrane‐enriched fractions bound almost three times as much DFB as the homogenate. Repetitive shorttime incubations (up to 3 h) of suspended cells with DFB resulted in a stepwise intracellular accumulation of DFB. Treatment of growing cells with DFB at high concentrations (50 μM) of DFB for longer periods (up to 7 days) resulted in elevated intracellular accumulation of DFB, which exceeded the binding capacity of the cell membranes and the aqueous solubility of DFB. These results indicate that the intracellular crystals detected by transmission electron microscopy are precipitated DFB. No metabolites or other chemically modified products of intracellular DFB were detected by high pressure liquid chromatography (HPLC) after a 7‐day incubation.

Differential Age Susceptibility of the Codling Moth (Lepidoptera: Tortricidae) to Chitin Synthesis Inhibitors and Thuringiensin

The concentration-response of Cydia pomonella eggs and first instars to chitin synthesis inhibitors diflubenzuron, triflumuron, and CGA 112913 (2,6-difluoro- N -[[[4-[3-chloro-5-trifluoromethyl-2-pyridinyloxy]-3,5-dichlorophenyl]amino]carbonyl]benzamide), and the beta-exotoxin thuringiensin was determined in laboratory bioassays. Phosalone, the insecticide currently used for codling moth control in walnuts, was included in the tests for comparative purposes. Eggs were topically treated and larvae were fed surface-treated synthetic diet. Diflubenzuron and triflumuron were most toxic to 0- to 2-day-old eggs, and CGA 112913 and thuringiensin were nontoxic. In contrast, CGA 112913 was the most toxic compound to first instars: LC50 was 300-fold lower than LC50 for eggs. Triflumuron and diflubenzuron were 6- and 50-fold less toxic to first instars than to eggs, respectively, but still within the same range of toxicity as phosalone. LC50 of thuringiensin against first instars was also similar to that of phosalone. To determine the most sensitive period in the egg stage to triflumuron, hatch rates of 0- to 2-day-old eggs were compared with 2- to 4-day-old eggs. Because mortality of treated 2- to 4-day-old eggs was significantly lower than mortality of 0- to 2-day-old eggs, the critical stage for chitin synthesis occurs between 0 and 2 days. If applied to eggs, timing of sprays may be important.