Hormonal control of seasonal-morph determination in the small copper butterfly, Lycaena phlaeas daimio Seitz.

Spring and summer morphs of Lycaena phlaeas daimio Seitz. are characterized by a wing colour of red and reddish brown, respectively.When newly ecdysed pupae destined to be summer or intermediate morphs (90 or 10%) by larval exposure to long days (long-day pupae) were decapitated or decerebrated, more than half of the operated pupae developed into intermediate and spring morphs (48 and 7%). But, in pupae destined to be spring, intermediate, or summer morphs (72, 22 or 4%) (short-day pupae) these operations did not produce any significant changes in the seasonal morph.Brains excised from newly ecdysed long-day pupae were transplanted into the abdomen of decapitated short-day pupae of the same age. The implants changed most recipients into summer and intermediate morphs (46 and 36%). However, when the brains of short-day pupae were used, no significant changes occurred in the seasonal morph.When long-day or short-day pupae were treated with 20-hydroxyecdysone just after pupation, they produced more reddish wings than those of the untreated or saline-treated controls. When the application was followed by chilling, already known to induce the reddish morph, the effects of both treatments are cumulative so that more reddish adults developed.The results indicate that the brain of long-day pupae secretes a factor causing the wing to be brownish. In the absence (or low titre) of the factor, most short-day pupae develop into spring or intermediate morphs. Furthermore, ecdysteroids make the wing more reddish, when applied to newly ecdysed pupae.     

Hormonal control of seasonal morph determination in the swallowtail butterfly, Papilio xuthus L. (Lepidoptera: Papilionidae)

Seasonal morphs (spring and sumer forms) of Papilio xuthus L. are determined coincidentally with diapause and non-diapause in pupae by larval exposure to short days and long days respectively. The neuroendocrine principle underlying seasonal-morph determination was studied using surgical operations in P. xuthus.When recipient 0-day old or chilled diapause pupae were joined to donor 0-day old non-diapause pupae, the recipients developed into summer or intermediate morphs. When the same kinds of recipients used above were joined to 0-day old or chilled diapause pupae, there were no significant effects on the adult morph. In contrast, recipient non-diapause pupae all developed into summer morphs, regardless of groups of the type of donors.Furthermore, when the brain of 5th-instar larvae, pharate pupae or pupae, predetermined to be diapause, was transplanted into the abdomen of 0-day old, 30-day old or chilled diapause pupae, the recipients developed into summer or intermediate morphs.The results indicate that the brain of non-diapause pupae secretes a humoral factor producing the summer morph. In non-diapause pupae, the factor may be secreted at about the stage of larval-pupal ecdysis coincidentally with that of prothoracicotropic hormone.     

Hormonal control of seasonal morphs by the timing of ecdysteroid release in Araschnia levana L. (Nymphalidae: Lepidoptera)

Araschnia levana L. occurs in two seasonal morphs. Larvae reared under short-day conditions become diapause pupae and emerge as red spring-morph butterflies. Long-day larvae become non-diapause pupae, which emerge as black and white summer morphs. Pupae reared under these different conditions were joined in parabiosis. Both underwent adult development without diapause and the long-day animals developed into the summer morph as normal. The morph of short-day animals depended on the time of parabiosis. When they were joined to fresh long-day pupae 1 day after their own pupation, summer morphs resulted. When parabiosis began 4 days after pupation or later, spring morphs resulted. Extirpation of the brain-corpora cardiaca-allata complex from long-day pupae affected neither non-diapause development nor the summer morph. Adult development could be prevented by removal of head and prothorax. When adult development was initiated in the remaining bodies by 20-hydroxyecdysone 14 days after pupation, the spring morph resulted.—Injection of 20-dydroxyecdysone into 3-day-old short-day pupae initiated adult development and led to the summer morph. Injections into 10-day-old short-day pupae led to the spring morph. The same was true in diapause pupae deprived of their brain-corpora cardiaca-allata complex. These results indicate that seasonal diphenism in A. levana is controlled only by the timing of ecdysteroid release, which initiates adult development. There is no direct influence of the brain on wing coloration.     

Effect of larval brain extracts derived from three swallowtail butterflies,Papilio helenus L., P. machaon L. and P. memnon L., on seasonal morph development of P. xuthus L. (Lepidoptera: Papilionidae)

Adults of the three papilionid butterflies, Papilio helenus L., Papilio machaon L. and Papilio memnon L., exhibit seasonal diphenism comprising spring and summer morphs. To elucidate the physiological mechanism underlying seasonal morph development in papilionid butterflies, we investigated whether a cerebral factor showing summer‐morph‐producing hormone (SMPH) activity is present in the brain of three Papilio species using an assay system with chilled male short‐day pupae of P. xuthus L. When 2% NaCl extracts derived from 20 larval brains of the three species were injected into abdomens of chilled male short‐day pupae of P. xuthus, all recipients destined to develop into spring‐morph adults developed into summer‐ and intermediate‐morph adults. On the other hand, all recipients injected with distilled water as a control developed into spring‐morph adults. These results indicate that a cerebral factor showing SMPH activity is present in the larval brain of the three Papilio species. Additionally, all recipients injected with 2% NaCl extracts derived from 20 adult brains of Bombyx mori L. also developed into summer‐ and intermediate‐morph adults. The results revealed that SMPH or a cerebral factor showing SMPH activity is widely distributed among lepidopteran insects.     

Separation of Bombyxin from a neuropeptide of Bombyx mori showing Summer-morph-producing Hormone (SMPH) activity in the Asian Comma Butterfly,Polygonia c-aureum L

A neuropeptide from brain-suboesophageal ganglion (Br-SG) complexes of the silkmoth, Bombyx mori, shows summer-morph-producing hormone (SMPH) activity in the Asian comma butterfly, P. c-aureum. The SMPH-active peptide was extracted and demonstrated to be almost the same molecular size as bombyxin (4-5kD), a nueropeptide which shows prothoracicotropic hormone (PTTH) activity when assayed in vitro with prothoracic glands (PGs) of 4th-instar B. mori larvae in vitro. A Sephadex G-50 fraction of 3-8kD molecules prepared from Br-SG complexes of B. mori adults was applied to CM-, SP-, DEAE- or QAE- Toyoperal columns at pH 5.6 (or pH 6.9). The SMPH-activity could be separated from the PTTH-activity (or bombyxin) by subjecting a SMPH- and PTTH-active preparation of B. mori to anion-exchange chromatography at pH 6.9. By reversed-phase HPLC following an anion-exchange chromatography, SMPH-activity was recovered in two fractions of 40-45% acetonitril. Results demonstrate that the B. mori peptide showing the SMPH-activity in P. c-aureum is a different molecule than bombyxin.     

Photoperiod- and temperature-dependent regulation of pupal beige/black polymorphism in the small copper butterfly, Lycaena phlaeas daimio Seitz

The small copper butterfly, Lycaena phlaeas daimio, has pupal beige/black polymorphism, the development of which is found to be controlled in an apparent association with the development of adult seasonal polymorphism (spring and summer morphs) by photoperiod and temperature in the larval stages. That is, the pupae of beige and black types developed under long-day and short-day conditions tend to develop into brown-winged and red-winged adults, respectively. In addition, a large proportion of long-day pharate pupae chilled at 4 degrees C for 5 days were observed to develop into pupae whose head-thoracic complexes and abdomens were judged to be of the black and intermediate types, respectively. They developed into adults with redder wings as compared to those obtained from unchilled pupae. The results indicate that the physiological mechanism underlying the photoperiodic control of the development of adult seasonal polymorphism may also play a significant role in the determination of pupal beige/black polymorphism in L. phlaeas daimio. Furthermore, cuticle melanization was found to be induced in the head-thoracic complexes of pupae by chilling of the pharate pupae. Melanization of pupal cuticle seems to occur in a close association with the development of reddish-winged adults.     

Effects of photoperiod and temperature on seasonal morph development and diapause egg oviposition in a bivoltine race (Daizo) of the silkmoth,Bombyx mori L

A bivoltine race (Daizo) of the silkmoth, Bombyx mori, has two types of adults which are classified on the basis of the number of dark-brown and brown scales distributed along the outer lines of the anterior wings. Development of these two types of adults is determined by photoperiod and temperature experienced during the embryonic and larval stages. The two types of adults are thought to be seasonal (summer and autumn) morphs. All adults developed under short days at low temperature (15 degrees C and 23 degrees ) were classified as summer morphs and oviposited only nondiapause eggs (ND-eggs), and those that developed under long days at high temperature (28 degrees C and 25 degrees C) were autumn morphs ovipositing only diapause eggs (D-eggs), only ND-eggs or a mixture of D- and ND-eggs. Larval exposure to long days at 25 degrees C shifted the wing pattern towards the autumn morphs and decreased the incidence of female adults ovipositing only D-eggs. Larval exposure to long days elicits an opposite effect on D-egg oviposition to that observed when long days are received only during the embryonic stage.     

Role of photoperiod and temperature in seasonal morph determination of the butterfly Eurema hecabe

ABSTRACT. Environmental cues affecting seasonal morph determination of the pierid butterfly Eurema hecabe L. were examined in laboratory experiments and in the field. A sexual difference exists in the photoperiodic response for seasonal morph determination: in short days at 25C the proportion of the autumn morph is higher in females than in males, and as the rearing temperature is decreased the proportion of autumn morphs increases in both sexes. A 5C drop in temperature during the final larval stadium induces 100% autumn morph production in short-day animals. This increase in autumn morph production induced by temperature decrease explains the seasonal morph change of this butterfly occurring in early autumn in the field. The role of this sex-linked photoperiodic response is discussed as a seasonal strategy of reproduction.     

Influence of generation and photoperiod on larval development of Lobesia botrana (Lepidoptera: Tortricidae)

The influence of generation (under field conditions) and photoperiod (under laboratory conditions) on Lobesia botrana larvae development was studied. Some larvae were collected during three annual generations in two grape-growing areas of northeastern Italy, and others were individually reared in the laboratory from egg to pupa on an artificial diet under two different photoperiod conditions (respectively, daylight 16 h/d [long day {LD}] and 14 h/d [short day {SD}]). The mandible lengths of collected larvae were measured and the data analyzed morphometrically to determine the number of larval instars. In the laboratory study, the number of larval moultings, the mandible length of each instar, the development time from hatching larva to pupa, and the pupal weight were considered. The measurement of mandible lengths of larvae collected in the field indicated the existence of five larval instars in all three annual generations, but the size of the two oldest larval instars was significantly higher for third-generation larvae than for the previous generations. Under laboratory conditions, the larvae usually exhibited five instars, but the mandible lengths of larvae and the pupa size were greater for individuals reared under SD. These also took a greater number of days to develop from hatching larvae to pupae. Because a larger size of the final larval instar occurs in individuals that produce diapausing pupae under SD in both the laboratory and the field, a positive association between larval size and the probability of surviving the winter can be inferred.     

Control of larval-pupal-adult molt in the moth Sesamia nonagrioides by juvenile hormone and ecdysteroids

Sesamia nonagrioides (Lepidoptera: Noctuidae) larvae reared under long day (LD; 16L:8D) conditions pupate after 5 or 6 larval instars, whereas under short day (SD; 12L:12D) conditions they undergo up to 12 additional molts before pupating. This extended period of repeated molting is maintained by high levels of juvenile hormone (JH). Previous work demonstrated that both LD and SD larvae decapitated in the 6th instar pupate but further development is halted. By contrast, about one-third of SD larvae from which only the brain has been removed, undergo first a larval molt, then pupate and subsequently developed to the adult stage. Debrained LD larvae molt to larvae exceptionally but regularly pupate and produce adults. Implanted brains may induce several larval molts in debrained recipient larvae irrespectively of the photoperiodic conditions. The results of present work demonstrate that the prothoracic glands (PGs) and the corpora allata (CA) of debrained larvae continue to produce ecdysteroids and JHs, respectively. PGs are active also in the decapitated larvae that lack JH, consistent with the paradigm that CA, which are absent in the decapitated larvae, are the only source of this hormone. Completion of the pupal-adult transformation in both LD and SD debrained insects demonstrates that brain is not crucial for the development of S. nonagrioides but is required for diapause maintenance. Application of JH to headless pupae induces molting, presumably by activating their PGs. It is likely that JH plays this role also in the induction of pupal-adult transformation in debrained insects. Application of the ecdysteroid agonist RH 2485 (methoxyfenozide) to headless pupae also elicits molting: newly secreted cuticle is in some cases thin and indifferent, in other cases it bears distinct pupal or adult features.     

The evolution of alternative morphs: density-dependent determination of larval colour dimorphism in a butterfly

Understanding the ultimate causes for the presence of polymorphisms within populations requires knowledge of how the expression of discrete morphs is regulated. In the present study, we explored the determination mechanism of a colour dimorphism in larvae of the butterfly Pararge xiphia (Satyrinae: Nymphalidae) with the ultimate aim of understanding its potential adaptive value. Last-instar larvae of P. xiphia develop into either a green or a brown morph, although all individuals are invariably green during the preceding three instars. A series of laboratory experiments reveal that morph development is strongly environmentally dependent and not the result of alternative alleles at one locus. Photoperiod, temperature, and in particular larval density, all influenced morph determination. The strong effect of a high larval density in inducing the brown morph parallels other known cases of density-dependent melanization in Lepidopteran larvae. Because melanization is often correlated with increased immune function, this type of determination mechanism is expected to be adaptive. However, the ecology and behaviour of P. xiphia larvae suggests that increased camouflage under high-density conditions may be an additional adaptive explanation. We conclude that the colour dimorphism of P. xiphia larvae is determined by a developmental threshold that is influenced both by heredity and by environmental conditions, and that selection for increased immune function and camouflage under high-density conditions may be responsible for maintaining the dimorphism.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 98 , 256–266.     

Morphogenesis and phenotypic divergence in two developmental morphs of Streblospio benedicti (Annelida,Spionidae)

Abstract. The morphology of marine invertebrate larvae is strongly correlated with egg size and larval feeding mode. Planktotrophic larvae typically have suites of morphological traits that support a planktonic, feeding life style, while lecithotrophic larvae often have larger, yolkier bodies, and in some cases, a reduced expression of larval traits. Poecilogonous species provide interesting cases for the analysis of early morphogenesis, as two morphs of larvae are produced by a single species. We compared morphogenesis in planktotrophic and lecithotrophic morphs of the poecilogonous annelid Streblospio benedicti from the trochophore stage through metamorphosis, using observations of individuals that were observed alive, with scanning electron microscopy, or in serial sections. Offspring of alternate developmental morphs of this species are well known to have divergent morphologies in terms of size, yolk content, and the presence of larval bristles. We found that some phenotypic differences between morphs occur as traits that are present in only one morph (e.g., larval bristles, bacillary cells on the prostomium and pygidium), but that much of the phenotypic divergence is based on heterochronic changes in the differentiation of shared traits (e.g., gut and coelom). Tissue and organ development are compared in both morphs in terms of their structure and ontogenetic change throughout early development and metamorphosis.     

The prediapause copulation and its significance in the butterflyEurema hecabe

In the pierid butterflyEurema hecabe, which shows seasonal polymorphism (summer and autumn morphs) and overwinters at adult stage, whether or not the prediapause copulation may be of usual occurrence and reproductively functional was examined. From the counts of the spermatophores carried by the females, it is clear that the prediapause copulation characteristically occurs in reproductively inactive females of the autumn corph. From behavioral observations in the field, mating partners of those females are mostly males of the summer morph rather than from the autumn morph. In autumn, males of the summer morph remained abundant and searched for females on the larval food plants. Furthermore, they mated frequently with autumn morph females. Autumn morph males seemed to be sexually less active or inactive before hibernation. Microscopic examinations of the spermathecae were made in mated autumn-morph females collected before and after hibernation. The results indicate that sperm is passed by the males at autumnal copulation. The sperm may be stored in the female reproductive tract and utilized for fertilization in spring. This supposition is strongly supported by field data; that is, once-mated autumn-morph females laid fertilized eggs in spring. Finally, physiological basis of the prediapause copulation, its adaptive signficance and the behavioral advantage inE. hecabe are discussed from the viewpoint of seasonal adaptation.     

The environmental and genetic control of seasonal polyphenism in larval color and its adaptive significance in a swallowtail butterfly

Seasonal polyphenism, in which different forms of a species are produced at different times of the year, is a common form of phenotypic plasticity among insects. Here I show that the production of dark fifth-instar caterpillars of the eastern black swallowtail butterfly, Papilio polyxenes, is a seasonal polyphenism, with larvae reared on autumnal conditions being significantly darker than larvae reared on midsummer conditions. Both rearing photoperiod and temperature were found to have individual and synergistic effects on larval darkness. Genetic analysis of variation among full-sibling families reared on combinations of two different temperatures and photoperiods is consistent with the hypothesis that variation in darkness is heritable. In addition, the genetic correlation in larval darkness across midsummer and autumnal environments is not different from zero, suggesting that differential gene expression is responsible for the increase in larval darkness in the autumn. The relatively dark autumnal form was found to have a higher body temperature in sunlight than did the lighter midsummer form, and small differences in temperature were found to increase larval growth rate. These results suggest that this genetically based seasonal polyphenism in larval color has evolved in part to increase larval growth rates in the autumn.     

Firing activity of "diapause hormone" producing cells in the male silkmoth,Bombyx mori

Diapause hormone (DH) originally identified to be a factor originating from neurosecretory cells in the suboesophageal ganglion acts on developing ovaries to produce diapause eggs in a female silkmoth, Bombyx mori. A male silkmoth has homologous neurosecretory cells, but little is known of the physiological nature of the cells and actions of their products. We examined the long-term firing activity of putative DH-producing neurosecretory cells and hormonal activity of their products in male pupae that had been experienced different environmental regimens for diapause induction. Firing activity patterns of male labial cells strongly depended on diapause types of pupae: cells in a diapause-type male were active throughout the pupal period, whereas the same cells in a non-diapause-type male were usually inactive during the early two-thirds of the pupal period. A male pupa with electrically active labial cells could induce diapause eggs in a female pupa connected parabiotically to that male. The firing activity of male neurosecretory cells and hormonal action of their products are qualitatively the same as in the female previously examined. We suggest that there is no evident sexual dimorphism in the physiological and biochemical nature of neurosecretory cells producing DH and the amidated peptide DH has different functions in a male.     

Effect of light quality and larval diet on morph induction in the polymorphic caterpillar Nemoria arizonaria (Lepidoptera: Geometridae)

The emerald moth Nemoria arizonaria (Geometridae) is bivoltine, with distinct broods of caterpillars hatching in the spring and summer. Caterpillars of the spring brood develop into mimics of oak catkins, while caterpillars of the summer brood develop into mimics of oak twigs. Previous rearing experiments showed that all caterpillars reared on oak catkins developed into catkin morphs, while all caterpillars reared on oak leaves developed into twig morphs, regardless of temperature or photoperiod. However, those previous rearing experiments did not control the colour of light perceived by the caterpillars independently of their dict. Since wavelengths of light perceived by some species of polymorphic caterpillars can influence their colour, it is possible that morph induction in Nemoria arizonaria is due to the characteristics of light reflected from yellow catkins or green leaves, rather than larval diet itself. The experiments reported here independently varied larval diet and light characteristics to determine if light quality is involved in morph induction. Only larval diet influenced morph induction, since all caterpillars reared on catkins developed into the catkin morph, and all caterpillars reared on oak leaves developed into the twig morph, regardless of whether they perceived yellow light, green light, or were raised in the dark.     

Effects of ecdysteroid agonist RH-2485 reveal interactions between ecdysteroids and juvenile hormones in the development of Sesamia nonagrioides

Larvae of Sesamia nonagrioides developing under long day (LD) conditions pupate in the 5th or 6th instar, whereas under the short day (SD) conditions, they undergo several supernumerary larval molts and are regarded as diapausing. The development in early larval instars occurs in the LD larvae at a moderate and in the SD larvae at a high juvenile hormone (JH) titer; ecdysteroid titer cycles similarly under both conditions. The transformation to pupa is initiated by a burst of ecdysteroids at undetectable JH levels, whereas extra larval molts in the diapausing larvae are associated with moderate JH titer and irregular rises of ecdysteroids. Application of 0.2 ppm RH-2485 to the diet of the 6th instar larvae promotes hormonal changes supporting metamorphosis in the LD larvae and slightly accelerates larval molts in the diapausing SD larvae. The 0.5- and 1-ppm doses revert these patterns of endocrine regulations to a mode typical for early larval instars. Particularly dramatic is a JH titer increase provoked within 24 h in the LD larvae. After the treatment, both the LD and SD larvae undergo a series of larval molts, suggesting that hormonal programming of the larval development has been stabilized. A few insects receiving 1 ppm RH-2485, and a high proportion of those fed with 5 ppm RH-2485, deposit two cuticles within a single apolysis and die.     

Phenotypic and genetic variation in emergence and development time of a trimorphic damselfly

Although colour polymorphisms in adult organisms of many taxa are often adaptive in the context of sexual selection or predation, genetic correlations between colour and other phenotypic traits expressed early in ontogeny could also play an important role in polymorphic systems. We studied phenotypic and genetic variation in development time among female colour morphs in the polymorphic damselfly Ischnura elegans in the field and by raising larvae in a common laboratory environment. In the field, the three different female morphs emerged at different times. Among laboratory-raised families, we found evidence of a significant correlation between maternal morph and larval development time in both sexes. This suggests that the phenotypic correlation between morph and emergence time in the field has a parallel in a genetic correlation between maternal colour and offspring development time. Maternal colour morph frequencies could thus potentially change as correlated responses to selection on larval emergence dates. The similar genetic correlation in male offspring suggests that sex-limitation in this system is incomplete, which may lead to an ontogenetic sexual conflict between selection for early male emergence (protandry) and emergence times associated with maternal morph.     

Kin recognition and cannibalism in polyphenic salamanders

We investigated kin discrimination among larvae of Arizona tigersalamanders (Ambystoma tigrinum nebulosum) which occur as "typical"morphs that feed mostly on invertebrate prey and occasionallyon conspecifics, and as "cannibal" morphs that feed primarilyon conspecifics. When housed with smaller larvae that differedin relatedness, both cannibals and typicals preferentially consumedless-related individuals. Cannibals ate typicals much quickerwhen the choice was between nonkin and siblings than when thechoice was between nonkin and cousins, indicating that cannibalscould distinguish different categories of relatives. Cannibalswere less likely to eat a larval sibling that was a cannibalmorph than a sibling that was a typical morph. Occluding animals'nares temporarily eliminated kin discrimination, implying thatolfaction is important in recognition. Larvae from differentsibships varied considerably in their ability to discriminatekin, and the greater the probability that a larva from a givensibship would develop into a cannibal morph, the more likelythe members of that sibship were to discriminate kin. Our resultsenable us to infer the functional significance of kin recognitionin this species and to develop an evolutionary model of themechanisms underlying the joint control of kin recognition andcannibalistic polyphenism.