环丙氨嗪对鸡粪中亮斑扁角水虻生长的影响及添加活性炭对环丙氨嗪的解毒作用

【目的】研究环丙氨嗪对鸡粪中亮斑扁角水虻Hermetia illucens幼虫生长发育的影响及添加活性炭对环丙氨嗪的解毒作用。【方法】使用含不同浓度(0, 0.1, 0.25, 0.5, 1, 2.5, 5, 10和15 mg/kg)环丙氨嗪的鸡粪分别饲养3, 4和5龄亮斑扁角水虻幼虫,统计幼虫的生长发育(体重和死亡率)变化;在鸡粪中添加不同浓度(0, 10, 30, 50, 100和150 g/kg)活性炭饲养亮斑扁角水虻3龄幼虫,测定活性炭对其生长发育的影响以及对环丙氨嗪的解毒作用。【结果】在用含一定浓度环丙氨嗪的鸡粪饲养后,亮斑扁角水虻3, 4和5龄幼虫均表现出体节伸长,停止进食,直至死亡。鸡粪中环丙氨嗪对亮斑扁角水虻3, 4和5龄幼虫的致死中浓度(medium lethal concentration, LC₅₀)值分别为0.18, 1.39和6.45 mg/kg。在鸡粪中添加量为30～100 g/kg的活性炭可以显著促进亮斑扁角水虻3龄幼虫的生长发育,并且能够解除环丙氨嗪对3龄幼虫的毒性,95%有效剂量(95%effective dose, ED     

Stage-dependent effects of 20-hydroxyecdysone on DNA synthesis of corpus allatum cells in the silkworm,Bombyx mori

DNA synthesis in cells of the corpus allata (CA) of the silkworm, Bombyx mori, was studied immunocytochemically after in vivo labeling with 5-bromo-2'-deoxyuridine (BrdU); developmental changes during the 3rd, 4th, and last larval instars and effects of 20-hydroxyecdysone treatment were examined. During both the 3rd and 4th larval instars, the number of DNA-synthesizing cells fluctuated, and relatively low levels were observed during the middle stages. On day 0 of the last larval instar, the number of DNA-synthesizing cells per gland was 9.2, which then increased on day 1 and remained at levels ranging from 12.9 and 16.9 cells per gland. A major peak level (28 BrdU-labeled cells per gland) occurred on day 8, two days after larvae entered the wandering stage. When last instar larvae were fed 20-hydroxyecdysone-supplemented mulberry leaves starting on day 0 or 1, the number of DNA-synthesizing cells dramatically decreased to very low levels and these low levels were maintained throughout the remainder of the instar. However, no effect was observed when last instar larvae were fed 20-hydroxyecdysone-supplemented mulberry leaves starting on day 3, indicating the stage-specific action of 20-hydroxyecdysone. The mechanism by which 20-hydroxyecdysone treatment inhibits DNA synthesis of CA cells was further examined by using continuous in vitro BrdU labeling for a 2-day incubation. It was found that the decrease in responsiveness of DNA synthesis of CA cells of 20-hydroxyecdysone-treated larvae to stimulation by growth factors from hemolymph may have been, at least in part, responsible for the indirect inhibitory effects of 20-hydroxyecdysone.     

Inhibition of growth and development of tobacco hornworm (Lepidoptera: Sphingidae) larvae by cyromazine

When fed to larvae of tobacco hornworm, Manduca sexta (L.), cyromazine strongly inhibited normal growth and development. Depending on the dose or period of feeding, symptoms were cuticular melanization, swellings in intersegmental regions, cuticular lesions, rupture of the body wall, and death. At greater than 10 ppm, cyromazine in the diet initiated symptoms in fifth instars, whereas greater than 20 ppm induced symptoms which led to death during the instar or at molting. The ratio of chitin to larval body weight in both cyromazine-exposed and control fifth instars increased slightly from ecdysis to 42 h. However, this ratio did not differ between the two groups, indicating that cyromazine had no immediate or direct effect on chitin production. When cyromazine was ingested by fifth instars, it was excreted rapidly, and a small amount (less than 5%) was metabolized to melamine. The amount of cyromazine found in the hemolymph remained relatively constant during the feeding period, whereas the amount present in the body wall increased with time and was localized in the KOH-soluble fraction. Cyromazine may inhibit growth or expansion of the body wall (or both) sufficiently to prevent normal internal growth, producing the observed symptoms and leading to abnormal development.     

Haemolymph ecdysteroid titre and epidermal cell commitment of the female southwestern corn borer larvae

The epidermal cell commitment (to pupation or formation of immaculate larvae) and related haemolymph ecdysteroid titres of the southwestern corn borer, Diatraea grandiosella were studied in both nondiapause-bound and diapause-bound last-instar female larvae. Cell commitment was estimated by examining the characteristics of new cuticle secreted in response to an injection of 20-hydroxyecdysone. Haemolymph ecdysteroid titres were determined by radioimmunoassay. Juvenile hormone effect on epidermal cell commitment was studied by applying a juvenile hormone mimic (ZR-515) to last-instar non-diapause-bound larvae and examining the resulting cuticle.In non-diapause-bound larvae, the epidermis of different body regions was committed to pupal development at different times. When pupal cuticular characteristics were evaluated by a scoring system, it appeared that the development of normal pupal cuticle is discontinuous. Three sudden increases in pupal characteristics were observed at 1.67, 2.67 and 3.67 days into the last-larval instar. Haemolymph ecdysteroid titre changes were correlated with the sudden increases in pupal characteristics. Peak ecdysteroid titres were found at 1.67, 2.33, and 3.33 days into the final instar. A fourth ecdysteroid peak (138.8 ng/ml of haemolymph) occurred in pharate pupae. In contrast, the commitment of diapause-bound larvae to produce immaculate integument was made in a fast and continuous fashion. Full commitment was made by 50% of the individuals 4 days (ca. first quarter) into the stadium. Haemolymph ecdysteroid titres fluctuated during the first 2 weeks of the stadium but no significant peaks were observed prior to pharate stage. An ecdysteroid peak (29.8 ng/ml of haemolymph) was identified in pharate immaculate larvae.Pupal development could be completely prevented in 26.7% of nondiapause-bound larvae as late as 4 days into the last instar by topical application of ZR-515. This indicates that the commitment to pupation as revealed by 20-hydroxyecdysone injection is reversible.     

Effects of ultralow doses of fenoxycarb on juvenile hormone–regulated physiological parameters in the silkworm,Bombyx mori L.

Effects of fenoxycarb at ultralow doses were investigated on juvenile hormone (JH)–regulated parameters in the silkworm, B. mori. Like JH, this non-terpenoid carbamate is able to induce permanent larvae in the last larval instar. However, whereas micrograms of JH are needed to produce this effect, only a few picograms of fenoxycarb are necessary to induce the same effect. The effects of fenoxycarb observed in this study were only visible from day 4 of the last larval instar—that is, when the JH titer has dropped to undetectable levels and JH-repressed physiological parameters would naturally be expressed. We observed that the permanent larvae induced with low doses of fenoxycarb (100 pg/larva) had no 20-hydroxyecdysone (20E) peak. Their prothoracic glands (Pgs) were completely inactive and very weakly sensitive to prothoracicotropic hormone (PTTH). Fenoxycarb at doses of 1 ng/larva also significantly inhibited silk gland growth and coloration, whereas carotenoid content of the hemolymph was maintained at high levels, which could reflect an inhibition of its uptake by the silk glands. Total hemolymph protein levels in last instar larvae were also depressed at these doses. So, it seems that low doses of fenoxycarb are sufficient to maintain in a juvenilized status the physiological parameters that are normally expressed when JH titer has declined. Moreover, from an endocrinological viewpoint, we demonstrated that the corpora allata (CA) are not necessary for fenoxycarb to induce those effects and discussed its possible mode of action. Arch. Insect Biochem. Physiol. 37:178–189, 1998. © 1998 Wiley-Liss, Inc.     

Candidate target mechanisms of the growth inhibitor cyromazine: studies of phenylalanine hydroxylase, puparial amino acids, and dihydrofolate reductase in dipteran insects

Cyromazine, an insect growth regulator, affects larval and pupal cuticles in dipterans and some other insects. The mode of action of this aminotriazine is not known yet, though it has been shown not to inhibit the synthesis of chitin and cuticular proteins. Cyromazine may, however, act on some step(s) of sclerotization of the cuticle. In the present study, we have analyzed the key enzyme for the production of sclerotization agents, phenylalanine hydroxylase (PAH), using the enzyme from Drosophila, a cyromazine-sensitive insect. PAH was studied in vitro with cyromazine and three biologically less active derivatives at concentrations ranging from 1 microM to 1 mM. None of the compounds did significantly affect PAH activity. Nor did cyromazine, fed to last instar larvae of Musca domestica, change the relative content of phenylalanine and tyrosine, or the total amount and profile of amino acids of puparial cuticles, which showed a larviform shape typical for cyromazine intoxication. Taken together, this study does not support the hypothesis that phenylalanine hydroxylase represents a target site of cyromazine. In additional studies, the conflicting results, as reported by others, on in vitro inhibition of dihydrofolate reductase (DHFR) by cyromazine were re-examined using the enzymes from larvae of the blowfly, Protophormia terraenovae, and from hen liver. There was no significant inhibition of either DHFR at 100 microM by cyromazine as well as by dicylanil, a pyrimidine analog that is biologically more active than cyromazine. In conclusion, the mode of action of cyromazine remains completely open.     

Long-term effects of Bacillus thuringiensis subsp. israelensis on Aedes aegypti.

Immediate (24 hours exposure) and long-term (until the emergence of the adults) effects of different doses of a primary powder of Bacillus thuringiensis subsp. israelensis (B.t.i.) against first and second instar Aedes aegypti larvae were monitored. The long-term effect was dose-dependent and was materialized by a prolongation of the preimaginal development and continuous cumulative mortality until the emergence of the adults. Mortality values were higher during the fourth larval instar and the pupal stage. Some of the larvae reaching the fourth instar were smaller in size and remained in this state a 2-4 times longer period than in the control and finally died as larvae or very small pupae. The long-term effect was more intense as the treatment was applied earlier during the larval development. The correlation of the immediate lethal effect with the late effect allows the evaluation of the total impact of a larvicidal treatment.     

Mode of action of the venom of the ectoparasitic wasp,Euplectrus comstockii, in causing developmental arrest in the European corn borer,Ostrinia nubilalis

Euplectrus comstockii Howard (Hymenoptera: Eulophidae), is an ectoparasitic, gregarious wasp which parasitizes the larval stage of several important lepidopteran pests. Parasitization of both natural and unnatural hosts prevents molting in the parasitized instar. Here we report the effect of wasp venom on the European corn borer (unnatural host), an important pest of corn and other vegetables. Venom collected from venom glands of adultE. comstockii, when injected intoO. nubilalis 5th instars, inhibited the growth rate, development and molting of the injected larvae. The observed effect on molting was dose and age dependent. When 3rd, 4th and 5th instarO. nubilalis were envenomated by adult wasps, the larvae also were developmentally arrested and failed to undergo a molt. However, 3rd and 4th instars underwent apolysis (separation of the epidermis from the old cuticle) and produced new cuticle. Fifth instars did not. A premolt hemolymph ecdysteroid peak was not observed in these experimental 5th instars, but injections of 20-hydroxy-ecdysone induced apolysis and new cuticle formation. Envenomated 4th instars (on becoming pharate 5th instars) exhibited a premolt hemolymph ecdysteroid peak. HPLC/RIA revealed that 20-hydroxyecdysone was present in the hemolymph of these pharate 5th instars. Thus, in the European corn borer, the mode of action of the venom depended upon the instar parasitized. Our results support the presence of a venom component(s) that, in 4th instar hosts, inhibited ecdysis, but did not prevent hemolymph ecdysteroid levels from increasing sufficiently to stimulate apolysis. In 5th instars, the same, or perhaps, a different component(s) ofE. comstockii venom prevented the synthesis/release of ecdysteroid by inhibiting a previously unknown molt-regulating physiological event that occurs between days 3 and 4 of the instar. Deceased     

Effects of brief exposures to low temperature on the development, longevity and fecundity of the grain aphid Sitobion auenae (Hemiptera: Aphididae)

First instar nymphs and adults of the grain aphid Sirobion auenae that had been reared at 10°C and 20°C over a number of generations, were cooled to -5°C and -10°C for 1 h and 6 h and returned to 20°C to assess the effects of brief exposures to low temperatures (cold-pulses) on their survival. rate of development, longevity and fecundity. A strong acclimation response was observed in first instar nymphs, with significantly less mortality in groups reared to 10°C compared to 20°C. Mean development time from first instar to adult was not significantly affected by low temperature exposure at the first nymphal stage. Longevity in all groups cooled as first instars was reduced by the sub-zero cold-pulses, and was also dependent on temperature and exposure time. Acclimated aphids survived longer than non-acclimated individuals. Reproductive rate, in terms of the number of nymphs born per aphid per day, was unaffected by cold stress applied at the first instar stage. Total fecundity was however reduced, being a function of the number and longevity of the survivors. Adult aphids were less cold hardy than nymphs; mortality was higher at -10°C than -5°C increasing with duration of exposure from 1 h to 6 h. Mean fecundity was reduced significantly in aphids cooled at the adult stage, the number of aphids born per day decreasing as the exposure period of the cold-pulse increased, suggesting that low temperature had affected embryogenesis. All the nymphs born to adults surviving exposure to -5°C for 6 h died within 48 h of birth, indicating that low temperature has a pre-natal effect on mortality.     

Mechanisms of midgut remodeling: juvenile hormone analog methoprene blocks midgut metamorphosis by modulating ecdysone action

In holometabolous insects such as mosquito, Aedes aegypti, midgut undergoes remodeling during metamorphosis. Insect metamorphosis is regulated by several hormones including juvenile hormone (JH) and 20-hydroxyecdysone (20E). The cellular and molecular events that occur during midgut remodeling were investigated by studying nuclear stained whole mounts and cross-sections of midguts and by monitoring the mRNA levels of genes involved in 20E action in methoprene-treated and untreated Ae. aegypti. We used JH analog, methoprene, to mimic JH action. In Ae. aegypti larvae, the programmed cell death (PCD) of larval midgut cells and the proliferation and differentiation of imaginal cells were initiated at about 36h after ecdysis to the 4th instar larval stage (AEFL) and were completed by 12h after ecdysis to the pupal stage (AEPS). In methoprene-treated larvae, the proliferation and differentiation of imaginal cells was initiated at 36h AEFL, but the PCD was initiated only after ecdysis to the pupal stage. However, the terminal events that occur for completion of PCD during pupal stage were blocked. As a result, the pupae developed from methoprene-treated larvae contained two midgut epithelial layers until they died during the pupal stage. Quantitative PCR analyses showed that methoprene affected midgut remodeling by modulating the expression of ecdysone receptor B, ultraspiracle A, broad complex, E93, ftz-f1, dronc and drice, the genes that are shown to play key roles in 20E action and PCD. Thus, JH analog, methoprene acts on Ae. aegypti by interfering with the expression of genes involved in 20E action resulting in a block in midgut remodeling and death during pupal stage.     

Effects of ingested phytoecdysteroids on the growth and development of two Lepidopterous larvae

The effects of ingested or injected 20-hydroxyecdysone on silkworm larvae (Bombyx mori) including death without moulting, death following completion of promoted moulting, death during promoted moulting (ecdysis inhibition) and inhibition in growth with and without effects on moulting, are dependent upon the concentration of exogenous hormone, the precise developmental stage of the treated larvae, and the duration of exposure to the exogenous ecdysteroid. Comparisons of 20-hydroxyecdysone with other phytoecdysteroids in the silkworm and pink bollworm, Pectinophora gossypiella, show a similar but more potent effect induced by ponasterone A, while cyasterone causes an ‘antiecdysone’ effect.     

Hormonal regulation of macromolecular synthesis in testes and accessory reproductive glands of Spodoptera litura during post-embryonic and adult development

In S. litura testicular growth during the last larval instar and early pupal stage is associated with significant increase in DNA, RNA and protein contents. DNA synthesis is stimulated by 20-hydroxyecdysone (20-HE) in the penultimate instar testes. 20-HE injection in ligated late last instars increases the testicular weight and protein content. Accessory reproductive gland (ARG) development takes place during the mid and late pupal stages. Protein synthesis in the pharate adult ARG is stimulated by 20-HE. Juvenile hormone has no effect on ARG protein synthesis.     

Nutritional and hormonal effects on storage proteins in the silkworm,Bombyx mori L.

Starvation of 48 h old fifth instar larvae depressed storage protein titres initially for 48 h but retained the levels comparable to control thereafter, possibly due to nutrients obtained during the 48 h feeding after fourth ecdysis. After an initial decline ligated larvae accumulated maximum storage proteins in haemolymph. This is because of inhibitory juvenile hormone titre at the basal level besides the appropriate release of 20-hydroxyecdysone from the ectopic source(s). Injection of methoprene (10 Μg/larva) repressed accumulation of storage proteins while 20-hydroxyecdysone (10 Μg/larva) increased the same. P-soyatose injection to starved and ligated larvae accelerated storage protein accumulation in haemolymph, signalling nutrient indispensability for initiation of storage protein synthesis at the appropriate time of last instar development inBombyx mori.     

In vitro bioassay for the effect of Ajuga reptans phytoecdysteroids on Trialeurodes vaporariorum larval development

A rapid and reliable in vitro bioassay method to test the effects of synthetic and natural compounds on the development of greenhouse whitefly Trialeurodes vaporariorum Westwood (Homoptera Aleyrodidae) is described.Leaf fragments of Nicotiana glauca Graham infested with eggs of whitefly were cultured under aseptic conditions on half-strength Murashige-Skoog medium. The development of the insect in vitro and in vivo were comparable except for a higher mortality rate in the first larval instar.Ecdysteroids from Ajuga reptans L. were applied to leaf fragments by immersion in aqueous solution of the compounds or by addition to the culture medium. A high mortality rate of first larval instars was detected with 29-norsengosterone and ajugalactone treatments. This effect was specially pronounced when the products were incorporated in the medium. A significant increase in mortality was also recorded in the pupal state with a 20-hydroxyecdysone treatment. This effect was never observed before, because all whitefly larvae on A. reptans leaves die in the first larval instar.     

Control of activity and regression of the silk glands in the last-larval instar of Galleria mellonella

The weight increase of silk glands in isolated larval abdomens is enhanced by brain implants which stimulate RNA and protein synthesis. It is proposed that, in intact last-instar larvae, a blood-borne factor from the brain promotes silk gland activity during the facultative feeding period. In the post-feeding larvae, some starving larvae and in isolated larval abdomens the silk glands regress. The regression is accelerated and the glands degenerate under the action of 20-hydroxyecdysone. This effect is not associated with an increase in protease and RN-ase activities. In the normal larvae, the protease activity increases gradually during the last instar to a maximum at the time of regression and drops in the degenerating glands. RN-ase activity is maintained at a high level in the fully active, regressing and degenerating glands The results indicate that regression and degeneration of the silk glands are caused by structural cell reorganization rather than by an appearance of cytolytic enzymes.     

几种杀虫剂对南美斑潜蝇及其寄生蜂潜蝇茧蜂的毒力

测定了阿维菌素、灭蝇胺、杀虫单、毒死蜱、灭多威、顺式氯氰菊酯6种杀虫剂对南美斑潜蝇2龄幼虫、成虫及其寄生蜂潜蝇茧蜂的室内毒力,并考察了灭蝇胺、阿维菌素和杀虫单对南美斑潜蝇的田间防效。在所研究浓度下,阿维菌素、灭蝇胺对2龄幼虫的毒力较高,但对南美斑潜蝇成虫和潜蝇茧蜂的毒力较低;杀虫单对南美斑潜蝇成虫、2龄幼虫和潜蝇茧蜂的毒力均较高;毒死蜱、灭多威、顺式氯氰菊酯对南美斑潜蝇2龄幼虫的毒力较低,而对潜蝇茧蜂的毒力较高。田间条件下,灭蝇胺、阿维菌素和杀虫单都对田间南美斑潜蝇具有较高防效。     

Progress of developmental programme during the last larval instar of Bombyx mori: Relationships with food intake,ecdysteroids and juvenile hormone

The progress of developmental programme in the epidermal cells of last instar larvae of Bombyx mori was determined by ecdysteroid injections in normal and in JH-treated larvae. To clarify the importance of food intake in the control of development, starved animals were also used.The instar begins with a period during which the larval programme is expressed: this occurs in the presence of 20-hydroxyecdysone. Epidermal cells can thereafter secrete pupal cuticle after ecdysteroid injection although the larval programme is normally still present. During the last period only pupal characters can be expressed either in normal or in 20-hydroxyecdysone-injected larvae.These different developmental phases are not correlated with obligatory and facultative feeding periods.Transition from the first to the second phases is correlated with the absence of JH effects on pupal genes. JH applications during the second period, however, prevent the expression of pupal characters after 20-hydroxyecdysone injection. Thus, during this period, the pupal programme is not stabilized. Cellular reprogramming itself occurs at the onset of the last developmental period and is probably under the control of ecdysteroids.     

Entomogenous fungus Nomuraea rileyi inhibits host insect molting by C22-oxidizing inactivation of hemolymph ecdysteroids

The entomogenous fungus Nomuraea rileyi reportedly secretes a proteinaceous substance inhibiting larval molt and metamorphosis in the silkworm Bombyx mori. We studied the possibility that N. rileyi controls B. mori development by inactivating hemolymph molting hormone, ecdysteroids. Incubation of ecdysone (E) and 20-hydroxyecdysone (20E) in fungal-conditioned medium resulted in their rapid modification into products with longer retention times in reverse-phase HPLC. Each modified product from E and 20E was purified by HPLC, and identified by NMR as 22-dehydroecdysone and 22-dehydro-20-hydroxyecdysone. Some other ecdysteroids with a hydroxyl group at position C22 were also modified. Injection of the fungal-conditioned medium into Bombyx mori larvae in the mid-4th instar inhibited larval molt but induced precocious pupal metamorphosis, and its injection into 5th instar larvae just after gut purge blocked pupal metamorphosis. In hemolymph of injected larvae, E and 20E disappeared and, in turn, 22-dehydroecdysone and 22-dehydro-20-hydroxyecdysone accumulated. These results indicate that N. rileyi secretes a specific enzyme that oxidizes the hydroxyl group at position C22 of hemolymph ecdysteroids and prevents molting in B. mori larvae.     

Evaluation of synergistic interactions between the Colorado potato beetle (Coleoptera: Chrysomelidae) pathogen Beauveria bassiana and the insecticides, imidacloprid, and cyromazine

Laboratory studies investigated the interaction between the fungal entomopathogen Beauveria bassiana (Balsamo) Vuillemin and sublethal doses of the insecticides imidacloprid and cyromazine when applied to larvae of the Colorado potato beetle, Leptinotarsa decemlineata (Say). When second instars were fed potato leaf discs treated with sublethal doses of imidacloprid and a range of doses of B. bassiana, a synergistic action was demonstrated. Similar results were observed when larvae were sprayed directly with B. bassiana conidia and immediately fed leaf discs treated with imidacloprid. No synergistic interaction was detected when larvae were fed leaf discs treated with sublethal doses ofimidacloprid 24 h after application of B. bassiana conidia to larvae. However, a synergistic interaction was detected when larvae were fed leaf discs treated with imidacloprid and sprayed with B. bassiana conidia 24 h later. Although sublethal doses of both imidacloprid and the triazine insect growth regulator (IGR) cyromazine prolonged the duration of the second instar, only imidacloprid interacted with B. bassiana to produce a synergistic response in larval mortality. In leaf consumption studies, the highest dose of B. bassiana tested promoted feeding in inoculated second instars. Feeding was inhibited when larvae were fed foliage treated with sublethal doses of imidacloprid and significantly reduced when fed foliage treated with a sublethal dose of cyromazine. Starvation of larvae for 24 h immediately after B. bassiana treatment produced a similar result to the combined treatment of B. bassiana and imidacloprid and increased the level of mycosis when compared with B. bassiana controls. Imidacloprid treatment affected neither the rate of germination of B. bassiana conidia on the insect cuticle nor the rate at which conidia were removed from the integument after application. The statistical analysis used to detect synergism and the possible role of starvation-induced stress factors underlying the observed synergistic interactions are discussed.