Haemolymph-ecdysteroid peaks occur in headless larvae of Calpodes ethlius

During the last-larval stadium of Calpodes ethlius, there is a critical period after which neck ligation no longer prevents pupation. Radioimmunoassay of haemolymph from larvae ligatured after this critical period shows that the ecdysteroid titre remains lower than normal for 3 days then rises to a prepupal peak, falls to a low level, and rises rapidly again close to the time of pupation. Hormone peaks resembling those found in normal larvae are, therefore, produced in the absence of the head. The slowly rising hormone titre seen in normal larvae prior to the prepupal peak is abolished by neck ligation, indicating that this phase of the titre retains dependence on the head after the critical period. This difference may account for the lack of intermoult wax and endocuticle secretion in neck-ligated larvae. It is concluded that peaks of haemolymph ecdysteroids can be generated at appropriate developmental stages in the absence of the head, whereas the slowly rising phase of haemolymph ecdysteroids cannot.     

Haemolymph ecdysteroid titres in diapause and nondiapause larvae of the European corn borer, Ostrinia nubilalis

The ecdysteroid titres of last-instar prediapausing, diapausing and nondiapausing larvae of Ostrinia nubilalis were determined by radioimmunoassay. In the nondiapause larvae a major peak of ecdysteroid activity preceded pupation by 24 h and continued through the pupal ecdysis. This peak was correlated with head and thorax critical periods as well as with changes in behaviour and physiology marking the transition from feeding larva to prepupa. Nondiapause larvae also displayed a rise in ecdysteroid titre during the feeding phase of development. This rise was approx one tenth that of the major peak and lasted 32 h. It was not correlated with any overt changes in larval physiology or behaviour. The diapause ecdysteroid profile was distinctive in that the levels measured were all lower than the lowest of the nondiapause curve. No peaks were observed in the diapause titres. Prepupal changes such as spinning and the cessation of feeding were not correlated with any increase in ecdysteroid levels. During diapause termination, under a long-day photoperiod, no increases in ecdysteroid titre were observed for the first 10 days. After 12 days individuals began to show ecdysteroids above the diapause levels. Pupation started after 16 long days.     

Cellular events in the prothoracic glands and ecdysteroid titres during the last-larval instar of Spodoptera littoralis

Changes in prothoracic gland morphology were correlated to developmental events and ecdysteroid titres (20-hydroxyecdysone equivalents) during the last-larval instar in Spodoptera littoralis. After ecdysis to the last-larval instar the haemolymph ecdysteroid titre remained at about 45 ng/ml, when the prothoracic glands appeared quiescent. The first signs of distinct gland activity, indicated by increased cell size and radial channel formation, were observed at about 12 h prior to the cessation of feeding (36 h after the last-larval moult), accompanied by a gradual increase in ecdysteroid titre to 110 ng/ml haemolymph, at the onset of metamorphosis. During this phase ecdysteroid titres remained at a constant level (140–210 ng/ml haemolymph) and prothoracic gland cellular activity was absent for a short period. The construction of pupation cells occurred when haemolymph ecdysteroids titres increased to 700 ng/ml. A rapid increase in ecdysteroids began on the fourth night (1600 ng/ml haemolymph) reaching a maximal level (4000 ng/ml haemolymph) at the beginning of the fourth day. In freshly moulted pupae a relatively high ecdysteroid titre (1100 ng/ml haemolymph) was still observed, although during a decrease to almost negligible levels. The increase in ecdysteroid level during the third and the fourth nights of the last-larval instar was correlated with the period when almost all the prothoracic gland cells showed signs of high activity. Neck-ligation experiments indicated the necessity of head factors for normal metamorphosis up to the second to third day of the instar. The possibility that the prothoracic glands are under prothoracicotropic hormone regulation at these times is discussed.     

Haemolymph ecdysteroid in the housefly,Musca domestica,during oögenesis and its relationship with vitellogenin levels

Ecdysteroid titre in the haemolymph of the housefly, Musca domestica, cycled during oögenesis and peaked at ～50 pg/μl during stages 5, 6 and 7. Levels of 10–20 pg/μl were found in houseflies with pre- and post-vitellogenic ovaries. Removal of the corpus allatum and corpus cardiacum complex resulted in low ecdysteroid levels (10 pg/μl). Ovariectomized flies also had lower ecdysteroid levels than the controls at 2 days (5 pg/μl) after emergence but not at 6 days (22 pg/μl). It is possible that the ecdysteroid peak that occurred during stages 5, 6 and 7 was produced by the ovaries because ovaries secreted and synthesized ecdysteroid in vitro. Endogenous haemolymph ecdysteroid levels had a linear correlation with the amount of vitellogenin that held for hormone concentrations of 5–43 pg/μl. Furthermore, the injection of 20-hydroxyecdysone at doses of 10 ng?1.0 μg/fly increased the amount of vitellogenin from 6 h to 12 h after injection; by 24 h, the vitellogenin returned to control levels. When 20-hydroxyecdysone was injected into ovariectomized flies, it was rapidly degraded and 96% was cleared from the haemolymph within 1 h.     

Haemolymph ecdysteroid titres during larval-adult development in Rhodnius prolixus: Correlations with moulting hormone action and brain neurosecretory cell activity

A haemolymph ecdysteroid titre of the fifth (last)-larval instar of the hemipteran, Rhodnius prolixus has been determined by radioimmunoassay. During the last-larval stadium the ecdysteroid titre increases from a negligible level in the unfed insect to a detectable level within minutes following a blood meal. The titre reaches a plateau of ～50–70 ng/ml at 3–4 hr and this level is maintained until day 5–6, the time of the head-critical period in Rhodnius. At the head-critical period the titre begins to increase again, this time dramatically, reaching a peak of ～ 3500 ng/ml at day 13. From day 14 to ecdysis (day 21) the titre declines to a low level, ～ 30 ng/ml. Basal levels of ecdysteroids, ～ 15 ng/ml, were detectable in young adult males and females. A survey of haemolymph volumes during the last-larval instar indicates that the changes in the ecdysteroid titre reflect changes in the rates of ecdysteroid synthesis, and not changes in haemolymph volume. Excretion of ecdysteroids varies systematically during the instar, suggesting that control of ecdysteroid excretion may be important in regulation of the haemolymph titre. Qualitative analysis of the haemolymph ecdysteroid RIA activity revealed the presence of only ecdysone and 20-hydroxy-ecdysone. For the large peak preceding larval-adult ecdysis, 20-hydroxy-ecdysone was the predominant hormone. These results indicate that there may be two periods of release of prothoracicotropic hormone (PTTH) from the brain in Rhodnius, one immediately following the blood meal and the second on day 5 or 6. The significance of these times of PTTH release is discussed in relation to classical evidence of the timing of moulting hormone action, the response of target tissues, and with more recent findings on the timing of release of neurosecretory material from the brain of Rhodnius during moulting.     

Changes in ecdysteroid and juvenile hormone levels in developing eggs of Bombyx mori

It is well-known that insect eggs can contain very high concentrations of ecdysteroids, which undergo drastic changes during embryogenesis. We found that this is equally valid for juvenile hormones. Three juvenile hormone-immunoreactive compounds were observed in developing Bombyx mori eggs. They were assumed to be juvenile hormones 1, 2 and 3 according to their retention time in HPLC. These hormones underwent drastic and sudden changes. In the space of one day their concentration was seen to rise rapidly from an undetectable level up to as high as 4 × 10^?6 micromoles per mg of eggs. Their presence was detected as early as the first day of embryonic development, as well as during the blastokinesis period (day 5 to day 9) and in late embryos (day 12 to day 14). Their relative concentrations varied greatly. On two occasions, day 1 and day 8, all three hormones were simultaneously present. Moreover, juvenile hormone 3 was present during the blastokinesis period, either alone or in combination with hormone 2. The latter was the only hormone present in late embryos, before hatching. Thus, with regard to both ecdysteroids (ecdysone and 20-hydroxyecdysone) and juvenile hormones, each day of embryonic development displayed a different hormonal pattern. These patterns undoubtedly constitute a “hormonal code” of embryogenesis control. While 20-hydroxyecdysone can be assumed to trigger cuticulogenesis in embryos as it does in larvae, the effects of the other hormones as well as their possible interactions are questionable.     

Divergent rectrix moult: the implications and conditions of moult sequence

Wing and tail morphology strongly affect flight performance which may consequently decline during feather moult due to the creation of feather gaps in the flight‐surface. Hence, the size and shape of moult‐related gaps may directly affect flight capacity. Here, I examined the divergent rectrix moult sequence compared to the more common distal moult sequence. In the divergent moult, the focus of rectrix moult is shifted from the tail centre (R₁; rectrices numbered distally from mid‐tail outward) to another rectrix (R₂ or R₃), and then rectrices are moulted bidirectionally, towards the tail centre and outwards. The result of this moult sequence is the splitting of the tail gap into multiple smaller gaps. Using a large moult database including 5669 individuals of 47 Western Palaearctic passerine species, I found evidence of divergent moult sequence for only seven species. Using comparative and experimental approaches, I found that the divergent rectrix sequence is correlated with higher moult speed and lower aerodynamic cost. Furthermore, the divergent rectrix sequence is more common among adults than juveniles. This work focused on the feather moult sequence – a seldom studied aspect of the avian life‐history. I propose that moult‐related aerodynamic costs may be an important evolutionary factor not only in moult speed, but also in moult sequence.     

Haemolymph ecdysteroid titres in diapause- and non-diapause-destined larvae and pupae of Sarcophaga argyrostoma

No differences were observed between the rates of development of larvae and pupae from diapause- and non-diapause-destined lines of Sarcophaga argyrostoma except that those destined for diapause have a longer post-feeding, wandering, larval phase associated with a lower haemolymph ecdysteroid titre, as measured by radioimmunoassay. Following pupariation, both cultures show a high haemolymph titre associated with larval/pupal apolysis. The developing culture displays an ecdysteroid peak at 72 h after pupariation which may be involved with pupal/adult apolysis and the initiation of pharate-adult development. This peak is reduced in the diapause-destined culture. Following the initiation of pharate adult development, there is a very large peak at 85–90 h. Those pupae entering diapause display very low titres as a result of the failure of the brain/prothoracic gland axis to release ecdysone. There are no quantitative or qualitative differences between the titres of specific ecdysteroids in the prepupae of the two lines as determined by reverse-phase high-performance liquid chromatography. A preliminary examination of the levels of free and conjugated ecdysteroids has provided the basis for proposing a mechanism of ecdysone metabolism in this insect.     

Relations between ecdysteroid levels and pupal development in the ecd-1 temperature-sensitive mutant of Drosophila melanogaster

Relations between ecdysteroid levels and pupal development were studied in the temperature-sensitive mutant ecd-1, shifted from 20°C to 29°C at different stages. Ecdysteroid titration, using radioimmunoassays, revealed that the mid-pupal peak of ecdysteroids was not affected when the shift was performed at pupariation although, in this condition, pupae died at emergence. A qualitative study of ecdysteroid content using HPLC failed to show the lack of any specific product. If the shift was performed 24 hr before pupariation, the ecdysteroid level decreased significantly while animals died before emergence but after a mid-pupal peak (around 75% of pupal life). However, double-shift experi-experiments showed that, even though a partial rescue of pupal development occurred (7% emergence), a normal mid-pupal peak was not restored. Furthermore, the temperature-sensitive periods for the decrease in the ecdysteroid mid-pupal peak and for the lack of emergence were not the same. Altogether, these results suggest that a decrease in ecdysteroids might not be the primary effect of the ecd-1 mutation, as the developmental anomalies observed at the end of pupariation seemed independent of the ecdysteroid levels.     

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.     

The occurrence of the second moult of Ascaris lumbricoides and Ascaris suum

Maung M. 1978. The occurrence of the second moult of Ascaris lumbricoides and Ascaris suum. International Journal for Parasitology 8: 371–378. Eggs of Ascaris lumbricoides and A. suum were cultured at 28°C and observed daily. Larvae were released by pressure, by artificial hatching with CO₂, and by natural hatching after infection of laboratory mice. The early stages of development in the egg were observed to comprise two moults, one occurring immediately after the other. Both moults were initiated within the egg, but the time of completion of the second moult varied considerably, and in some instances was not completed until the larvae reached the liver of experimentally infected animals.     

Variations in ecdysteroid levels in 5th instar larvae of Schistocerca gregaria in gregarious and solitary phases

Moulting hormone levels in whole bodies of 5th instar S. gregaria were determined throughout the 5th larval instar using gas chromatography with electron capture detection of the derivatized hormone. No differences were found between hormone levels in insects in the gregarious and solitary phase. No evidence for the existence of polar ecdysteroid conjugates used for storage or transport of these hormones was found. Approximately one tenth of the 20-hydroxyecdysone observed in the gregarious insects was detected in the faeces, almost equally divided between free hormone and polar conjugates of it.     

Characteristics of development and variations in ecdysteroid levels in Clitumnus embryos deprived of their cephalic endocrine glands

Microsurgical suppression of presumptive endocrine areas in very young embryos of Clitumnus resulted in an arrest in development, which occurred at different stages according to the operation. In most cases however. the duration of embryonic life was not reduced significanly. The repercussions of each operation type on the ecdysteroid content of the embryos were determined using a radioimmunoassay technique.Consideration of the experimental results provides some indication on the part played by each endocrine formation during the last part of embryonic development that is between dorsal closure and hatching time. Our experiments offer original evidence that the corpora allata of the embyro are of utmost importance at the stage VII_3t, in initiating the normal expression of larval characteristics.     

Haemolymph levels of juvenile hormone,ecdysteroids and protein during the last two larval instars of Locusta migratoria

The radioimmunoassay of ecdysteroids and juvenile hormone has enabled us to relate hormone levels to haemolymph protein concentrations and weight increase during the 4th and 5th instar of the migratory locust. The two hormones are never present in high concentrations in the blood simultaneously. The levels of ecdysteroids are high on the 5th day during the 4th larval stage: they show a small peak on the 3rd day, and then a large peak on the 8th day during the 5th instar. JHI-immunoreactive substances are high during the first 4 days of the 4th instar, and during the first 5 hr during the 5th instar. Protein concentrations in the haemolymph begin to rise when ecdysteroid levels increase during stage 4, and immediately after the small peak (on day 3) in the 4th stage larva. The rise in protein levels is correlated with an increase in weight.     

Haemolymph lipase activity in the American cockroach, Periplaneta americana

The haemolymph of the cockroach, Periplaneta americana, has been shown to possess pronounced lipolytic activity. The enzyme may be considered a ‘true’ lipase because of its ability to hydrolyse long chain fatty acid glycerides. The enzyme exhibits dual optima at pH 4·9 and 7·0 and is inhibited by fluoride ion. Lipase activity in the intact cockroach is unaffected by the corpus cardiacum hypolipemic factor; however, prolonged periods of starvation result in a significant decline in enzyme activity.     

Drosophila Fragile X Protein controls cellular proliferation by regulating cbl levels in the ovary

FMRP is an RNA binding protein linked to the most common form of inherited mental retardation, Fragile X syndrome (FraX). In addition to severe cognitive deficits, FraX etiology includes postpubescent macroorchidism, which is thought to result from overproliferation. Using a Drosophila FraX model, we show that FMRP controls germline proliferation during oogenesis. dFmr1 null ovaries contain egg chambers with both fewer and supranumerary germ cells. The mutant germaria contain a significantly increased number of cyclin E and PhosphoHistone H3 positive cells, suggesting that loss of FMRP leads to defects in cell cycle progression. BrdU incorporation and flow cytometry data suggest that, in addition to proliferation, germline endoreplication and ploidy are also affected by the loss of FMRP during ovary development. Here we report that FMRP controls the levels of cbl mRNA in the ovary and that reducing cbl gene dosage by half rescues the dFmr1 oogenesis phenotypes. These data support a model whereby FMRP controls germline proliferation by regulating the expression of cbl in the developing ovary.     

Haemolymph protein patterns during the spinning stage and metamorphosis of Rhynchosciara americana

An acrylamide gel electrophoretical analysis of the haemolymph proteins of R. americana was carried out at different stages of development. In mature larvae there are about 14 haemolymph protein fractions from which one stains heavily and two others faintly for lipoprotein, while three fractions stain for glycoprotein. The haemolymph protein fraction with Rm 0·25 decreases remarkably in mass during spinning, while the others decrease to a lesser extent. The protein fractions could be used in cocoon spinning since previous work suggests that haemolymph proteins are a major pool of cocoon protein precursors. The finding of a protein fraction in the salivary glands with an electrophoretical mobility similar to that of the haemolymph fraction with Rm 0·25 reinforces our hypothesis.     

Haemolymph ecdysteroid levels in female tsetse fly Glossina fuscipes (Diptera) during the first reproductive cycle: A comparison between virgin and mated females

The fluctuations in haemolymph ecdysteroid levels were recorded by radioimmunoassay during the first two ovarian cycles of the tsetse fly, Glossina fuscipes. During the first ovarian cycle, the patterns of ecdysteroid levels are similar in virgin and mated females. In virgins, the first oöcyte is not ovulated and ecdysteroid levels during the second vitellogenesis remain low. In mated females, vitellogenesis of the second oöcyte is concomitant with growth of the first larva in the mother's uterus and ecdysteroid levels are higher than during the first cycle. Two ecdysteroid increases were recorded in mated females and not in virgins. One coincides with vitellogenesis of the second oöcyte and the other occurs at the end of pregnancy and finishes after larviposition. The roles of ecdysteroids in the regulation of vitellogenesis, ovulation, pregnancy and larviposition are discussed.     

Haemolymph ecdysone concentrations in Hyalophora cecropia pupae,dauer pupae and adults

Haemolymph ecdysone concentrations were determined by radioimmunoassay in diapausing pupae, pharate adults, adults, and chilled dauer pupae. The concentration in diapausing pupae after 6 months chilling (5.35 pg/μl) increased dramatically after 3 days at 27°C (>200 pg/μl) and then decreased to low levels in adult females (1.63 pg/μl). In adult males ecdysone was undetectable in all except one animal. Dauer pupae showed a decrease from 6.1 to 1.7 pg/μl 1 day after being transferred from 6 to 27°C. Over a 3-day period the value increased to 3.19 pg/μl and remained constant for more than a year. These results suggest that diapausing pupae with and without brain neurosecretory cells maintain a low concentration of ecdysone in the haemolymph.     

Social aggregation of adult Schistocerca gregaria and Locusta migratoria migratorioides in relation to the final moult and ageing

Laboratory experiments, were carried out on nymphs and adults of the desert locust, Schistocerca gregaria, and the African migratory locust, Locusta migratoria migratorioides, in view of the field observation that locusts become temporarily less gregarious after the final moult. The experiments showed no reduction in grouping after the final moult but did show a reduction in grouping in mature Schistocerca. There was little change in the numbers of locusts found in small groups but a dramatic reduction in the number found in large groups.