The role of juvenile hormone in immune function and pheromone production trade-offs: a test of the immunocompetence handicap principle

The immunocompetence handicap hypothesis postulates that secondary sexual traits are honest signals of mate quality because the hormones (e.g. testosterone) needed to develop secondary sexual traits have immunosuppressive effects. The best support for predictions arising from the immunocompetence handicap hypothesis so far comes from studies of insects, although they lack male-specific hormones such as testosterone. In our previous studies, we found that female mealworm beetles prefer pheromones of immunocompetent males. Here, we tested how juvenile hormone (JH) affects male investment in secondary sexual characteristics and immune functions in the mealworm beetle, Tenebrio molitor. We injected male mealworm beetles with JH (type III) and found that injection increased the attractiveness of male pheromones but simultaneously suppressed immune functions (phenoloxidase activity and encapsulation). Our results suggest that JH, which is involved in the control of reproduction and morphogenesis, also plays a central role in the regulation of a trade-off between the immune system and sexual advertisement in insects. Thus, the results reflect a general mechanism by which the immunocompetence handicap hypothesis may work in insects.     

Juvenile hormone mediates a trade-off between primary and secondary sexual traits in stalk-eyed flies

Trade-offs between developing body parts may contribute to variation in allometric scaling relationships in a variety of taxa. Experimental evidence indicates that both circulating levels of juvenile hormone (JH) and sensitivities of developing body parts to JH can influence morphology in polyphenic insects. However, the extent to which JH may regulate both the development of traits that scale continuously with body size and trade-offs between these traits is largely unknown. Here, I present evidence that the JH analog methoprene applied to final instar larvae of a stalk-eyed fly (Cyrtodiopsis dalmanni) can induce males to produce larger eye-stalks relative to their body size. Examination of testis growth, sperm transfer, and egg maturation indicates that JH induces a trade-off between eye-span and gonad development in adult males, but not females. Age at sexual maturity was unaffected by larval JH applications to either sex. Collectively, these results are consistent with JH-mediated allocation of resources to eye-span at the expense of testes, and indicate potential costs for the production of an exaggerated trait.     

Ontogenetic colour change signals sexual maturity in a non-territorial damselfly

Conspicuous colouration increases male reproductive success through female preferences and/or male–male competition. Despite the advantages of conspicuous colouration, inconspicuous male morphs can exist simultaneously in a population due to genetic diversity, condition dependence or developmental constraints. We are interested in explaining the male dichromatism in Xanthagrion erythroneurum damselflies. We reared these damselflies in outdoor insectaries under natural conditions and showed that this species undergoes ontogenetic colour changes. The younger males are yellow and change colour to red 6–7 days after their emergence. We took red and yellow male reflectance spectra and found that red males are brighter than yellow males. Next, we aimed to determine whether ontogenetic colour change signals sexual maturity with field observations and laboratory experiments. Our field observational data showed that red males are in higher abundance in the breeding territory, and they have a higher mating frequency than yellow males. We confirmed these field observations by enclosing a red and a yellow male with two females and found that yellow males do not mate in presence of red males. To determine whether colour change signals sexual maturity, we measured mating success of males before and after colour changes by enclosing a single male at different age (day 3-day 7) and colour (yellow, intermediate and red) with a single female in a mating cage. Males did not mate when yellow but the same male mated after it changed colour to red, suggesting the ontogenetic colour change signals sexual maturity in this species. Our study shows that male dichromatism can be age-dependent and ontogenetic colour change can signal age and sexual readiness in non-territorial insects.     

Novel roles for the corpus allatum hormone in the cost of sexual interactions in the linden bug Pyrrhocoris apterus

The cost of sexual interactions, usually expressed as a reduction of life-span, is a fundamental but poorly understood aspect of life. According to a widely accepted view, a rise in the “pro-aging” juvenile hormone (JH) might contribute to the decrease of life span caused by sexual interactions. We tested this hypothesis using the linden bug Pyrrhocoris apterus by removing the corpus allatum (CA), the source of JH. If JH is causally involved in the cost of sexual interactions, then the absence of CA (JH) should decrease the negative effect of sexual interactions on survival. As expected, ablating the CA significantly prolonged life-span of both virgin females and virgin males. Mated insects of both sexes lived significantly shorter than virgins. However, contrary to prediction, the decrease of life span by sexual interactions was similar in control and CA-ablated males, and was even enhanced in CA-ablated females. Another unexpected finding was that males paired with CA-ablated females lived almost as long as virgin males and significantly longer than did males paired with control females, although ablating the female CA did not cause any decrease in mating activity. On the other hand, females paired with CA-ablated males lived only slightly longer than did females paired with control males. These results highlight several important points. (1) In both genders, the negative effect of sexual interactions on insect's survival is not mediated by the insect's own CA. (2) The male CA has only minor effect on female survival, while (3) the female CA (JH) is principally responsible for the sex-induced reduction in the male survival.     

Development of a real-time PCR assay for measurement of yellow protein mRNA transcription in the desert locust Schistocerca gregaria: a basis for isolation of a peptidergic regulatory factor

A major unresolved issue in insect endocrinology concerns the question of whether or not insects have sex hormones. Conclusive evidence in favor of the presence of such hormones awaits the establishment of appropriate bioassays in males. The cuticle of sexually mature males of the desert locust Schistocerca gregaria turns yellow in gregarious conditions only. Neither females nor isolated males ever turn yellow. The yellowing is due to the deposition in the cuticle of a male-specific Yellow Protein (YP), of which the amino acid sequence is known. In this paper, we describe the partial cloning of the cDNA encoding this Yellow Protein. The tissue distribution and temporal expression of the YP-mRNA is studied in detail using RT-PCR. Furthermore, an RT-PCR based bioassay was developed, which may serve as a reliable tool to help identify the hormones controlling the yellowing process. In addition to juvenile hormone, we have shown that a factor present in the brain-corpora cardiaca is involved in the yellow coloration, as injection of an extract induces the expression of YP-mRNA in isolated gregarious males.     

Influence of sexual organs on corpora allata biosynthetic activity in Locusta migratoria

ABSTRACT. Using an in vitro radiochemical assay, juvenile hormone (JH) biosynthesis by corpora allata was screened during the first 2 weeks of adult maturation in both male and female migratory locusts, Locusta migratoria migratorioides (R. & F.). The patterns of change of glandular activities in the sexes were different, but the overall rates of JH production were similar. Testisectomy did not appear to affect corpus allatum activity, male sexual behaviour, or yellow coloration. Removal of the testes together with the seminal vesicle and the accessory glands resulted in a disturbance of the pattern of JH biosynthesis, but had no effect on sexual behaviour or coloration. These observations are discussed in relation to available evidence on JH-mediated sexual events in the adult.     

Effect of juvenile hormone on senescence in males with terminal investment

Senescence, a decline in survival and reproductive prospects with age, is controlled by hormones. In insects, juvenile hormone (JH) is involved in senescence with captive individuals, but its effect under natural conditions is unknown. We have addressed this gap by increasing JH levels in young and old wild males of the damselfly Hetaerina americana. We assessed survival in males that were treated with a JH analogue (methoprene), which is known to promote sexual activity, and an immune challenge, which is known to promote terminal investment in reproduction in the studied species. We replicated the same procedure in captivity (to control for environmental variation), where males were deprived of any activity or food. We expected old males to show the lowest survival after being treated with JH and immune‐challenged, because the effect of terminal investment on senescence would be exacerbated by JH. However, this should be the case for wild animals, but not for captive animals, as the effects of JH and immune challenge should lead to an increase in high energetic‐demanding activities only occurring in the wild. Old animals died sooner compared with young animals in both the wild and captivity, confirming that males are subject to senescence. In wild but not captive animals, JH decreased survival in young males and increased it in old males, confirming that JH is sensitive to the environment when shaping animal senescence. Immune challenge had no effect on survival, suggesting no effect of terminal investment on senescence. Additionally, contrary to the expected effects of terminal investment, with an immune challenge, recapture rates increased in young males and decreased in old males. Our results show that male senescence in the wild is mediated by JH and that terminal investment does not cause senescence. One explanation is that animals undergoing senescence and terminal investment modify their feeding behaviour to compensate for their physiological state.     

Endocrine manipulations and ontogenesis of male sexual behaviour in Locusta: studies on azadirachtin-induced over-aged nymphs

ABSTRACT. Dose-response relations of the moult-inhibitory and lethal effects of azadirachtin (AZA) injected into fifth and fourth instar hoppers of Locusta migratoria migratorioides are presented. The optimal dose for obtaining maximum number of long-surviving over-aged nymphs is 1.6 and 1.0 μg of AZA per insect for young fifth and fourth instar nymphs, respectively. Over-aged male nymphs are capable of exhibiting sexual behaviour. The mating behaviour of fifth instar over-aged male nymphs, and of chemically allatectomized but otherwise similar over-aged male nymphs (obtained by combined precocene treatment and injection of AZA), with and without injections of exogenous Juvenile Hormone III (JH), is investigated quantitatively; the results are compared with those obtained for normal, chemically allatectomized, and chemically allatectomized plus JH-injected adult males. Over-aged fifth instar male nymphs show a low intensity of mating behaviour and in chemically allatectomized ones this intensity is even lower. Injections of JH intensify the mating behaviour of chemically allatectomized fifth instar over-aged male nymphs in a dose-dependent way, but less markedly so than in chemically allatectomized adult males. We conclude that the intensity of male mating behaviour of the over-aged nymphs is subnormal and their response to the mating behaviour intensifying effect of the JH is weaker than that of adult males. The relations between endocrine factors and ethological ontogenesis of male sexual behaviour in locusts are discussed.     

JH III production, titers and degradation in relation to reproduction in male and female Anthonomus grandis

Juvenile hormone (JH) is necessary for the production of vitellogenin (Vg) in the boll weevil, Anthonomus grandis. Occurrence of Vg in this species is typically restricted to reproductively competent females, and is not detected in untreated males. However, the JH analog, methoprene stimulates Vg production in intact males and in the isolated abdomens of both male and female boll weevils (where in each case no Vg is detected without treatment), suggesting that males are competent to produce Vg but are normally not stimulated to do so. Preliminary work indicating that male boll weevil corpora allata (CA) produced little or no JH in vitro suggested that failure of males to produce Vg might be due to very low JH levels compared to females. This study re-examines the question of JH in male boll weevils by determining in vitro production of JH III by male CA during the first 10 days after adult emergence, determining hemolymph JH esterase activity during this same time period and hemolymph JH III titers in adults of both sexes. We also re-examine the ability of isolated male abdomens to produce Vg in response to hormonal stimulation, analyzing the effect of a wide range of methoprene and JH III dosages. Results indicate that male A. grandis have circulating JH titers and JH production similar to females. JH esterase activity is slightly but significantly higher in males than females. Vg production by isolated abdomens of both sexes after stimulation with methoprene or JH III was confirmed. Dose response studies indicated that high levels of methoprene were less effective than intermediate doses in stimulating Vg synthesis in both sexes. We conclude that the sexually dimorphic effect of JH on Vg synthesis is not due to differences in JH production or differences in JH titer between the sexes.     

Juvenile hormone and photoperiodically controlled polymorphism in Aphis fabae: Postnatal effects on presumptive gynoparae

Long days (short nights) (LD 16:8) and high temperatures (> 15°C) have an apterizing effect on the short day (LD 12:12) induced, presumptive gynopara of Aphis fabae. Transfer of presumptive gynoparae to long days (15°C) or to 25°C (short days) at varying times during postnatal development demonstrate that the adult form is determined by the second day of the second instar, i.e. 5 days after birth at 15°C. Transfer on day 1 induces maximum apterization with the proportion of aphids affected decreasing with age at transfer.Apterization induced by long days immediately after birth can, to some extent, be cancelled by return to short days but only up to day 4. Thus long days are morphogenetically more potent than short days at the beginning of larval development. At temperatures above 15°C the proportion of aphids apterized increases almost linearly.Apterized insects can be distinguished from juvenilized insects in the fifth-instar. Topical application of juvenile hormone (JH) induces both apterization and juvenilization of presumptive gynoparae but at different times during larval development, JH treatment during the early-instars promotes apterization but induces little juvenilization, whereas maximum juvenilization, without apterization, is produced by middle-instar treatment. The apterizing effects of JH are, thus, not due to its neotenic action.The response profile of JH-induced apterization is similar to that observed with long days and 25°C. It is suggested that such conditions increase endogenous JH levels in A. fabae. The three naturally occurring JH's differ in activity in the order JH I > JH II > JH III. Both long-day and JH-apterized insects switch from the normal ovipara production of the adult gynopara to vivipara production.     

Age-dependent plasticity of sex pheromone response in the moth, Agrotis ipsilon: Combined effects of octopamine and juvenile hormone

Male moths use sex pheromones to find their mating partners. In the moth, Agrotis ipsilon, the behavioral response and the neuron sensitivity within the primary olfactory centre, the antennal lobe (AL), to sex pheromone increase with age and juvenile hormone (JH) biosynthesis. By manipulating the JH level, we previously showed that JH controls this age-dependent neuronal plasticity, and that its effects are slow (within 2 days). We hypothesized that the hormonal effect might be indirect, and one neuromodulator candidate, which might serve as a mediator, is octopamine (OA). Here, we studied the effects of OA and an OA receptor antagonist, mianserin, on behavioral and AL neuron responses of mature and immature males during stimulation with sex pheromone. Our results indicate that, although OA injections enhanced the behavioral pheromone response in mature males, OA had no significant effect on behavior in immature males. However, mianserin injections decreased the behavioral response in mature males. AL neuron sensitivity increased after OA treatment in immature males, and decreased after mianserin treatment in mature males. Determination of OA levels in ALs of immature and mature males did not reveal any difference. To study the possible interactive effects of JH and OA, the behavioral pheromone response was analyzed in JH-deprived mature males injected with OA, and in immature males injected with fenoxycarb, a JH agonist, and mianserin. Results show that both JH and OA are necessary to elicit a behavioral response of A. ipsilon males to sex pheromone.     

Juvenile hormone acts at embryonic molts and induces the nymphal cuticle in the direct-developing cricket

During embryogenesis of hemimetabolous insects, the sesquiterpenoid hormone, juvenile hormone (JH), appears late in embryogenesis coincident with formation of the first nymphal cuticle. We tested the role of embryonic JH by treating cricket embryos with JH III, or the JH-mimic (JHM) pyriproxifen, during early embryogenesis. We found two discrete windows of JH sensitivity. The first occurs during the formation of the first (E1) embryonic cuticle. Treatment with JHM prior to this molt produced small embryos that failed to complete the movements of katatrepsis. Embryos treated after the E1 molt but before the second embryonic (pronymphal) molt completed katatrepsis but then failed to complete dorsal closure and precociously formed nymphal, rather than pronymphal characters. This second sensitivity window was further assessed by treating embryos with low doses of JH III prior to the pronymphal molt. With low doses, mosaic cuticles were formed, bearing features of both the pronymphal and nymphal stages. The nymphal characters varied in their sensitivity to JH III, due at least in part to differences in the timing of their sensitivity windows. Unexpectedly, many of the JH III-treated embryos with mosaic and precocious nymphal cuticles made a second nymphal cuticle and successfully hatched. JH treatment also affected the growth of the embryos. By focusing on the developing limb, we found that the effect of JH upon growth was asymmetric, with distal segments more affected than proximal ones, but this was not reflected in misexpression of Distal-less or Bric-a-brac, which are involved in proximal-distal patterning of the limb.Edited by P. Simpson     

Endocrine manipulations and ontogenesis of male sexual behaviour in Locusta: studies on precocene-induced early adultiforms

ABSTRACT. Fourth- and third-instar permanent adultiforms of Locusta migratoria migratorioides (R.&F.) were produced by applying 7-ethoxy-precocene II (precocene III) to hoppers in earlier stages. The mating behaviour of male adultiforms and the effects of injected exogenous juvenile hormone III (JH) on this behaviour were investigated quantitatively and the results were compared to those obtained for normal adult males and for adult males chemically allatectomized by precocene III. Fourth- and third-instar adultiform males exhibited sexual behaviour and injections of JH intensified this behaviour in a dose-dependent manner, though the number of repeated injections in which a cumulative dose is administered was found to be not less important than the amount of the cumulative dose itself. Huge repeated doses (6×144=864 μg) of JH temporarily induced normal intensity of mating behaviour in fourth-instar male adultiforms. The same doses of JH intensified mating behaviour also in third-instar adultiform males, but less markedly so than in fourth-instar ones. For restoring temporarily normal intensity of mating behaviour in chemically allatectomized but morphogenetically normal adult males, much smaller doses (2×36=72, or 2×72=144μg) of JH were sufficient. In contrast, even huge doses of JH were unable to induce mating behaviour in normal (no precocene-treated) male hoppers. Thus, the ethological ontogenesis (='ethogenesis') of the male's mating behaviour, including the response of the system to JH, is clearly accelerated by precocious metamorphosis. However, this is not a simple 'all or none' effect, because adultiforms in earlier stadia exhibit less intense male mating behaviour and more limited response to JH than adultiforms in later stadia or adults.     

Sex pheromone and visual trap interactions in mate location strategies and aggregation by host-alternating aphids in the field

Abstract. Field observations were made on the responses of males and gynoparae of three host-alternating aphid species, the blackberry-cereal aphid, Sitobion fragariae (Walker), the bird cherry-oat aphid, Rhopalosiphum padi (L.) and the damson-hop aphid, Phorodon humuli (Schrank) to species-specific sex pheromones released from transparent and coloured water traps.Pheromone traps caught significantly more males than did control traps without pheromone, whereas transparent, light green, yellow and orange traps caught most insects.Measurements of the distance over which sex pheromones function indicated that male P.humuli detect the pheromone 2–6 m from the source and can fly upwind to a source in wind speeds of 0.7 m s^-1.In all three species significantly more gynoparae were caught in pheromone traps than in control traps, suggesting that pheromone released by adult sexual females may assist late-flying gynoparae to locate a suitable host plant on which to deposit their progeny.The response is relatively stronger for males than gynoparae, but the pheromones appear to act as both sex and aggregation pheromones.     

Nuclear development in locust fat body: the influence of juvenile hormone on inclusion bodies and the nuclear matrix

The hormonal induction of vitellogenesis in insects and in oviparous vertebrates are prime models of gene regulation in eukaryotes. In vertebrates the process is under estrogenic control and normally confined to females, although males can be artificially induced. In locust in contrast, juvenile hormone (JH) is central to fat body development in both males and females, yet the response is strongly sex limited not only for vitellogenin production but also in terms of total protein, DNA and RNA synthesis and nuclear ploidy levels. To differentiate further possible sex and/or JH related developmental aspects in locusts, large-scale nuclear events were examined during normal adult maturation and in animals treated with antiallatropins and JH analogs. Fat body nuclei undergo extensive restructuring during normal development in both sexes. This included progressive nuclear enlargement, accompanied by extensive proliferation of nuclear matrix components and elaboration of complex inclusion bodies (NB). The isolated protein matrix was unusually complex relative to similar structures from vertebrates and the NB were firmly anchored to it. Although matrix proteins were qualitatively similar to those from other sources, as assessed by SDS polyacrylamide gel electrophoresis, several major matrix polypeptides, including lamins A and B, and components greater than 150 kD, fluctuated quantitatively during development and in concert with nuclear enlargement. The number and morphology of the NB were unrelated to sex, but increased in direct proportion to absolute nuclear volumes. All changes were more pronounced in females, where higher ploidy levels, larger nuclei and correspondingly more internal matrix elements occurred. Suppression of JH production by precocene prevented all foregoing nuclear changes, but re-exposure to methoprene rapidly induced normal development. The results are compared to analogous nuclear changes in steroid responsive vertebrate tissues.     

Broad specifies pupal development and mediates the 'status quo' action of juvenile hormone on the pupal-adult transformation in Drosophila and Manduca

The understanding of the molecular basis of the endocrine control of insect metamorphosis has been hampered by the profound differences in responses of the Lepidoptera and the Diptera to juvenile hormone (JH). In both Manduca and Drosophila, the broad (br) gene is expressed in the epidermis during the formation of the pupa, but not during adult differentiation. Misexpression of BR-Z1 during either a larval or an adult molt of Drosophila suppressed stage-specific cuticle genes and activated pupal cuticle genes, showing that br is a major specifier of the pupal stage. Treatment with a JH mimic at the onset of the adult molt causes br re-expression and the formation of a second pupal cuticle in Manduca, but only in the abdomen of DROSOPHILA: Expression of the BR isoforms during adult development of Drosophila suppressed bristle and hair formation when induced early or redirected cuticle production toward the pupal program when induced late. Expression of BR-Z1 at both of these times mimicked the effect of JH application but, unlike JH, it caused production of a new pupal cuticle on the head and thorax as well as on the abdomen. Consequently, the 'status quo' action of JH on the pupal-adult transformation is mediated by the JH-induced re-expression of BR.