Application Of Methoprene Research Articles

Synthesis and Insect Growth Regulatory Activity of 1,5-Disubstituted Imidazoles with Non-terpene Chains.

1-Isobutyl-5-(4-phenoxyphenyl)imidazole (KK-98), an inhibitor of juvenile hormone (JH) biosynthesis in the cockroach, and related imidazole compounds were evaluated against silkworm, Bombyx mori, for their activity to induce precocious metamorphosis. KK-98 induced precocious metamorphosis in the 4th instar larvae at high doses. Replacement of the 4-phenoxy group by a 3-phenoxy or 3-benzyloxy group on the benzene ring increased the activity. Among this series of compounds, 5-(3-benzyloxyphenyl)-1-isopropylimidazole (8) showed the highest activity. The induction of precocious metamorphosis by compound 8 was rescued by the simultaneous application of methoprene, a JH minie. When newly molted 3rd instar larvae were treated with a high dose of compound 8, a few larvae formed larval-pupal intermediates in the 3rd instar stage, which has not been formed by treating of any other imidazoles so far.

Developmental expression and hormonal regulation of a desiccation stress protein in Tenebrio molitor

In a previous study it has been reported that dsp28 is induced during desiccation in Tenebrio larvae. During that study it was observed that in non-stressed larvae the concentration of dsp28 in hemolymph drops dramatically just prior to pupation. These results suggested that control of dsp28 synthesis is subject to environmental as well as hormonal cues. This study identifies a site of synthesis as fat body, as dsp28 was secreted into the medium during in vitro incubation of larval fat bodies. Using immunoelectrophoresis to determine protein concentration a developmental profile showing changes in levels of dsp28 in hemolymph during larval, pupal and adult stages of Tenebrio molitor was established. The concentration of dsp28 in larval hemolymph dropped from 4 to 5 mg/ml to 1.5 mg/ml just prior to pupation. This lower level was maintained until adults emerged when the concentration of dsp28 rose to prepupal levels again. Hormonal regulation is suggested since application of methoprene to newly-emerged pupae resulted in an increased incorporation of radiolabeled cysteine into dsp28.

Stimulated synthesis of the female‐specific storage protein in male larvae of the silkworm Bombyx mori treated with juvenile hormone analog

AbstractApplication of methoprene to fourth (penultimate) instar larvae of the silkworm Bombyx mori induced the appearance of the feeding dauer larvae at the fifth (last) instar and prevented pupal metamorphosis. Methoprene also increased the protein concentrations of hemolymph last instar larvae by preventing sequestration of storage proteins by the fat body. Usually, the female‐specific storage protein 1 (SP1)* disappears from the male hemolymph at the time of the last larval instar. However, exposure of male larvae to methoprene at the penultimate instar enhanced the accumulation of SP1 in the hemolymph. The SP1 accumulated in males did not differ in molecular weight and immunoreactivity from the SP1 produced in female larvae. Both sexes of fourth instar larvae allatectomized on day 1 instantly accumulated SP1 in the hemolymph, and methoprene application after allatectomy suppressed the hemolymph accumulation of the SP1. In contrast, if allatectomy was carried out at a later stage of the fourth larval instar, SP1 concentration in hemolymph of fifth instar larvae did not increase, suggesting the different juvenile hormone action for regulation of SP1 synthesis in the penultimate instar larvae of silkworms.

Ploidy levels and DNA synthesis in fat body cells of the adult mosquito, Aedes aegypti: The role of juvenile hormone

AbstractAdult females of the mosquito Aedes aegypti showed two cycles of DNA replication in the fat body based on microspectrophotometric measurement of changes in nuclear DNA. The first cycle began after emergence and resulted in 80% of diploid fat body cells becoming tetraploid and 20% becoming octoploid by the end of the third day. The second replication cycle occurred 48–72 h after a blood meal and resulted in an increase in octoploid nuclei to 67% Topical application of juvenile hormone or methoprene to abdomens isolated at emergence stimulated an increase in ploidy levels above that normally seen in situ. Synthesis of DNA, estimated by incorporation of injected [3H]‐thymidine, rose after emergence and remained high for 2 days. Synthesis increased again after a blood meal, reached a peak by 6 h, and returned to low levels by 24 h after the meal. The timing of DNA synthesis and a measurable increase in ploidy were temporally separated. The ploidy increase, but not DNA synthesis, was correlated with increases in juvenile hormone levels.

Ecdysteroid fluctuations in adult Drosophila melanogaster caused by elimination of pupal reserves and synthesis by early vitellogenic ovarian follicles

Radioimmunoassay (RIA) of whole body extracts of Drosophila melanogaster males and females demonstrates that at eclosion all individuals contain high levels of ecdysteroid. Highly polar ecdysteroids (presumably metabolites) in the meconium represent approximately half of the total ecdysteroid RIA-activity present at this time and are subsequently eliminated. Ecdysteroids remaining after the elimination of the meconium are also highly polar as shown by reverse-phase high pressure liquid chromatography (RP-HPLC). The amount of ecdysteroid RIA-activity found in whole body extracts declines in both sexes until 18 h post-eclosion when levels begin to increase in the female and drop to undetectable levels in the male. In the female the ovaries are the major source of ecdysteroid. The increase in whole body ecdysteroid in the female coincides with the initiation of ovarian ecdysteroid production and accumulation. Topical application of methoprene, a juvenile hormone (JH) analog, stimulates ovarian ecdysteroid synthesis in apterous-ts 78j (ap ts 78j) , a temperature-sensitive juvenile hormone-deficient mutant, corroborating previous results suggesting a role of juvenile hormone in ovarian ecdysteroid production. Stage 8–9 follicles, whose development is juvenile hormone dependent, are shown to be the most active in ecdysteroid production. The regulatory potential of these stages is discussed.

Precocene II treatment inhibits terminal oöcyte development but not vitellogenin synthesis and release in the black blowfly, Phormia regina Meigen

An antiallatal agent, precocene II. was used to assess the effect of the corpus allatum on vitellogenin synthesis/release and oöcyte development of the adult black blowfly, Phormia regina. Oöcyte development was greatly retarded by a treatment of precocene II given topically within 12 h after emergence. A single dose of 20 or 40 μg of precocene II per fly inhibited oöcyte development whereas a treatment given to flies more than 12 h after emergence was less effective. The precocene-inhibited oöcyte development could be reversed by a single treatment of methoprene—a juvenile hormone analogue. Scanning electron microscopy showed that the volume of the corpus allatum of precocene-treated females was significantly smaller than that of untreated females. Radiochemical assay showed that the juvenile hormone biosynthesis/release ( in vitro) by the corpus allatum of precocene-treated flies was significantly reduced but not stopped. In precocene-treated females, haemolymph protein and vitellogenin accumulated after a liver-meal, while ovarian soluble protein and vitellin content remained low. Application of methoprene lowered both the amount of haemolymph protein and vitellogenin and increased the quantities of the ovarian soluble protein and vitellin. Our data demonstrate that the major function of the hormone is to regulate the uptake of vitellogenin by developing oöcytes.

Interaction between juvenile hormone, 20-hydroxyecdysone, the corpus cardiacum-allatum complex, and the ovaries in regulating vitellogenin levels in the housefly, Musca domestica

This study examines the interaction between the ovaries, corpus cardiacum-allatum complex, juvenile hormone, and 20-hydroxyecdysone to either induce or regulate vitellogenin production in the housefly Musca domestica. All of the unoperated control flies contained vitellogenin and these levels were increased by the application of methoprene, a juvenile hormone analogue or 20-hydroxyecdysone. Flies without the corpus cardiacum-allatum complex had low vitellogenin levels, but these were restored to control amounts by treatment with methoprene, but not 20-hydroxyecdysone. Ovariectomized flies showed a steady increase in vitellogenin levels and only the treatment with 20-hydroxyecdysone increased the amounts of vitellogenin further. Flies without either the corpus cardiacum-allatum complex or the ovaries had undetectable amounts of vitellogenin, but treatment with methoprene gave a control pattern of vitellogenin, while 20-hydroxyecdysone treatment resulted in trace amounts of vitellogenin. In these studies only treatment with 20-hydroxyecdysone elevated vitellogenin to control levels if the corpus cardiacum-allatum complex was present. Treatment with methoprene elevated vitellogenin to control levels in flies without the corpus cardiacum-allatum complex. When ovariectomized insects without the corpus allatum-cardiacum complex were treated with 0.1 μg methoprene and 0.05 or 0.5 μg 20-hydroxyecdysone, vitellogenin levels were increased to control amounts, but neither hormone alone at these doses induced the appearance of vitellogenin. Thus it appears that the production of control levels of vitellogenin requires both juvenile hormone and 20-hydroxyecdysone.

Brachypterizing effect of group rearing, juvenile hormone III and methoprene in the wing-dimorphic cricket, Gryllus rubens

The effects of group rearing, topical application of juvenile hormone III, and topical application of methoprene on winglength development were studied in the wing-dimorphic cricket, Gryllus rubens. Group rearing had a strong brachypterizing effect on both an outbred polymorphic stock recently initiated from field-collected crickets and on a laboratory stock nearly pure-breeding for long wings. Transfer experiments demonstrated that group versus individual rearing during at least the first third of nymphal life exerts virtually no influence on the development of long versus short wings. Development could be redirected from long wings to short wings by transferring crickets from individual to group rearing as late as the last stadium. Topical application of juvenile hormone III during both the penultimate and last stadia and methoprene during the penultimate stadium also redirected development from long to short wings. Methoprene applied during the last stadium produced juvenile-adult intermediates. Both the brachypterizing and juvenilizing effect of methoprene occurred at a concentration one to two orders of magnitude lower than the lowest brachypterizing concentration of juvenile hormone. The results suggest that juvenile hormone is a morph-regulating hormone in G. rubens and provide the foundation for future physiological studies of sensitive periods in winglength development.

Hormonal control of cuticular melanization in the common cutworm, Spodoptera litura

The penultimate-instar larvae of Spodoptera litura raised under crowded conditions deposited melanized cuticle, which could be labelled by injected [ 14C]dopamine, while those reared in isolation deposited non-melanized cuticle. In crowded culture, melanization-and-reddish-colouration hormone was secreted between 6 and 9 h after head-capsule slippage, followed by melanization about 4 h later. Extirpation of the suboesophageal ganglion from the crowded larvae totally blocked melanization, while removal of the brain allowed 50% of the melanization as after denervation between the two organs. This suggests that the melanization-and-reddish-colouration hormone is secreted from the suboesophageal ganglion, and the brain regulates its secretion from that organ. Injection of melanization-and-reddish-colouration hormone of Bombyx mori into an isolated abdomen induced melanization, while this treatment combined with methoprene blocked it. Application of methoprene to crowded larvae up to 2 h before head-capsule slippage greatly reduced the black pigmentation. Allatectomy of the larvae reared in isolation could induce full melanization. These results indicate that a decrease in juvenile hormone titre at around the time of head-capsule slippage is essential for exhibiting the action of melanization-and-reddish-colouration hormone.

Enhanced production of aggregation pheromones in four stored-product coleopterans feeding on methoprene-treated oats

Production of aggregation pheromones by maleOryzaephilus surinamensis, O. mercator, Cryptolestes ferrugineus, andTribolium castaneum was enhanced by feeding on methoprene-treated oats, implicating juvenile hormone in control of pheromone production. Methoprene application to control insects in stored food products may cause enhanced pheromone production by these insects, thus drawing additional beetles into the treated product.

Differential timing of pupal commitment in various tissues of the silkworm, Bombyx mori

To determine the timing of the onset of pupal commitment of various tissues after the loss of juvenile hormone, penultimate-instar Bombyx mori larvae were allatectomized either 1 or 2 days after ecdysis, then the 0.1 μg of juvenile hormone analogue, methoprene, was applied at various times thereafter. At the next moult, the larval or pupal state of the imaginal discs, epidermis, silk glands and midgut was assessed. Spinning behaviour was also observed. Each tissue showed its own characteristic time of pupal commitment beginning with the silk glands at 22 h after allatectomy and ending with the mandibles at 84 h. Measurement of the haemolymph ecdysteroid at 3-h intervals revealed no increase in the low intermoult level until about 12 h after the methoprene application, then it increased to a moulting maximum by 48 h.

Developmental profiles of the mRNAs for Manduca arylphorin and two other storage proteins during the final larval instar of Manduca sexta

When fat body mRNA from the tobacco hornworm larva, Manduca sexta, was translated in a rabbit reticulocyte lysate system, three major polypeptides were found, each having a different developmental profile. One mRNA coded for a 74 kilodalton (K) polypeptide doublet precipitated by an antibody to the arylphorin (manducin). This mRNA was present only during the intermolt feeding phase of the penultimate and the final larval instars. Its appearance 16–24 hr after larval ecdysis was dependent upon the incoming nutrient supply and independent of the juvenile hormone (JH) level. Immunoblots of proteins of the fat body, epidermis, and cuticle revealed the presence of arylphorin in all three tissues. Additionally, several small polypeptides that cross-reacted with the arylphorin antibody were found in the fat body during and up to 24 hr after the last larval molt and in the tanning pupal cuticle. The larval epidermis was also found to contain a small amount of arylphorin mRNA. At the time of the JH decline prior to the onset of metamorphosis, a female-specific mRNA coding for a 79 K translation product appeared. In allatectomized larvae this mRNA was detectable earlier, and its appearance in intact larvae was prevented by application of methoprene, indicating that JH regulates its appearance. At wandering a new mRNA that also codes for a 79 K polypeptide appeared in both sexes and was the major messenger present during the prepupal stage. Neither it nor the female-specific mRNA were translatable after pupal ecdysis.

Effect of dietary ecdysterone on protein ingestion and copulatory behaviour of the blowfly, Sarcophaga bullata

ABSTRACT. In Sarcophaga bullata and other blowflies, the low haemolymph ecdysteroid titre is responsible for the inability of males to synthesize vitellogenin, although they have the genes coding for this protein. The high moulting hormone titre in females is of crucial importance for vitellogenesis. In this context we investigated whether the difference in protein ingestion between the sexes might also be related to differences in ecdysteroid titre. Using a two‐choice system, we observed that upon addition of ecdysterone to the food, protein ingestion increased in males. Topical application of methoprene also had a stimulating effect. Prolonged treatment with ecdysterone reduced the copulatory behaviour of males.

Laboratory and Field Trials of Soil Applications of Methoprene and Other Insecticides for Control of the Mediterranean Fruit Fly (Diptera: Tephritidae)1

The juvenile hormone analogue methoprene was shown to prevent adult eclosion in laboratory tests against Ceratitis capirata (Wiedemann). When used as a soil treatment in field trials, methoprene and methoxychlor were not effective. Our field trials demonstrated that diazinon was highly effective and that chlorpyrifos was only partially effective in preventing adult emergence from the soil.

Soldier determination in Pheidole bicarinata: Effect of methoprene on caste and size within castes

Topical application of methoprene to final-instar larvae of the ant Pheidole bicarinata can induce soldier development. Soldier induction takes place if methoprene levels are above a soldier-determining threshold during a critical period of juvenile hormone-sensitivity that occurs during about days 4–6 of the final instar. Furthermore, the amount of exogenous methoprene applied affects the timing of metamorphosis and the adult size in both the minor worker and soldier castes. When larvae that receive methoprene treatment become minor workers these are always larger than acetone-treated controls. In larvae that become soldiers, growth and timing of metamorphosis vary with the dose of methoprene, but in a more complex way. A high dose of methoprene produces a metamorphic delay and large soldiers. However, the lowest effective dose for soldier induction produces early metamorphosis and small adults. On the basis of these results, we have expanded our model of a mechanism by which juvenile hormone could control determination of worker castes in Pheidole bicarinata.

Effect of Methoprene on the Fecundity and Fertility of the Codling Moth, Cydia pomonella12

Female larvae of Cydia pomonella (L.) (Lepidoptera: Tortricidae) received a single topical application of a juvenile hormone mimic methoprene, at various intervals during the fifth (last) stadium. Adult survival rates were >75% when newly ecdysed, last-stadium larvae were treated with 0.01 to 1.0 μg of methoprene; however, survivorship decreased in populations treated later in that instar. The fecundity of surviving adults was reduced by 0.01 μg of methoprene applied to day 2 and day 6 5th instars.

Hormonal control of uric acid storage in the fat body during last-larval instar of Manduca sexta

In the last-larval instar of the tobacco hornworm, Manduca sexta, a switch from excretion of uric acid to storage in the fat body occurs during transition from the feeding to the wandering stage. Neuroendocrine control of this change from excretion to storage was demonstrated by neck-ligation experiments with synchronously reared larvae. Results indicate that a neurohormone is released from the head 24–30 hr before the initiation of wandering and coincident with the first release of ecdysone that initiates metamorphosis. Direct involvement of the moulting hormone was shown by the effects of multiple injections of 20-hydroxyecdysone into the abdomen of larvae that had been ligated before the release of hormone. Urate levels in the fat body were 20- to 100-fold higher from hormone-injected larvae as from saline inject controls. Topically applied juvenile hormone or methoprene reversed the 20-hydroxyecdysone-induced storage of urate. Increased levels of uric acid in the haemolymph during pupal development result from the presence of juvenile hormone, and the abrupt decrease in uric acid concentration in the haemolymph just prior to pupal ecdysis results from a decreased titre of juvenile hormone. Applications of methoprene prevented the decrease in uric acid levels in the haemolymph.

Diapause Termination, Gonoactivity, and Differentiation of Host-Seeking Behavior from Blood-Feeding Behavior in Hibernating Culex Tarsalis (Diptera: Culicidae)

Diapausing Culex tarsalis were collected from mine tunnels in Boulder Co., Colorado, during late September and early October 1980 and studied in the laboratory. Exposure to a long photophase (L:D 15:9) resulted in diapause termination in essentially all mosquitoes by day 7 at 25 °C. Warming alone (at 25 °C) for up to 8 days did not terminate diapause in a significant proportion of females maintained at typical autumn day lengths. Diapause was terminated in Cx. tarsalis by topical application of methoprene, a juvenile hormone (JH) mimic, suggesting that the physiological pathway for diapause termination involves secretion of JH by a reactivated corpus allatum. Host seeking and blood feeding were shown to represent discrete stages in the biting cycle of Cx. tarsalis , and these were differentiated in diapausing females by varying the size of the feeding chamber. Diapausing females could be induced to take a blood meal only when the host-seeking step in the biting cycle was bypassed by placing the females in close proximity to a host. Gonotrophic dissociation occurred in a significant proportion of females only when 3 conditions were met: females had to be in a state of diapause when fed, and fed females had to be incubated at a cool temperature (15 °C) and under short-day conditions during the time of blood meal digestion. The demonstration that gonotrophic dissociation can occur under experimental conditions probably has no relation to what occurs in nature. Warm Indian-summer conditions notwithstanding, typically short autumn day lengths are sufficient to suppress host-seeking behavior in diapausing populations and host seeking is a requisite for blood feeding in nature.

Susceptibility of Sitotroga cerealella (Oliv.) (Lepidoptera: Gelechiidae) to two insect juvenile hormone analogues

The effect of topical application of the insect juvenile hormone analogues methoprene (isopropyl-11-methoxy-3,7-11-trimethyl dodeca-2,4-dienoate) and hydroprene (ethyl-3,7,11-trimethyl dodeca-2,4-dienoate) on last instar larvae and newly-formed pupae of Sitotroga cerealella was examined. Topical application of methoprene at 10 μg/pupa to maize grains containing pupae resulted in 94% adult emergence whereas the same treatment applied to pupae removed from the grains reduced adult emergence to 45%. At 100 μg methoprene/pupa, adult emergence was 91% when grains containing pupae were treated and 32% when pupae were treated after removal from grains. Hydroprene (10 μg/pupa) was more effective than methoprene against insects both inside grains and after removal from grains, giving adult emergence of 48% and 8% respectively. At 100 μg/pupa, hydroprene completely prevented adult emergence when applied to insects removed from the grain. After treatment of the pupal stage with 10 or 100 μg methoprene per insect, adult males produced the same numbers of spermatophores as untreated males and the treatment did not affect either the number or fertility of eggs deposited by adult females.

Fluoromevalonate acts as an inhibitor of insect juvenile hormone biosynthesis

The potential of analogues of insect juvenile hormones (JHs) for pest control has been demonstrated by the application of methoprene for the selective control of certain insects, including several mosquito species, the cigarette beetle and the hornfly1. Compounds that impede JH production could offer a potentially more attractive method of control because accelerated metamorphosis would shorten the lifetime of the larvae, the stage that usually does maximum damage to the host plant. Of the few anti-juvenile hormone (AJH) agents described2–5, only precocenes evoke all the required biological responses, but they affect a few insect taxa of minor economic importance. Their activity is thought to result from cytotoxicity6,7 in the corpora allata. In view of similarities between the biosynthesis of the carbon skeletons of insect JHs8,9 and farnesyl pyrophosphate (a precursor of steroids and other terpenoids10), compounds that inhibit the early steps in the biosynthesis of cholesterol might be expected to suppress JHs. Although this concept has already been explored without success11,12, we now report the inhibition of juvenile hormone biosynthesis (AJH activity, see Table 1 legend) by a fluorinated mevalonate tetrahydro-4-fluoromethyl-4-hydroxy-2H-pyran-2-one, FMev, Fig. 1) which until now was known only to be hypocholesterolaemic13–15.