X-ray investigation of gas bubble formation, and water loss in tsetse fly pupae

ABSTRACT. Using a microfocal X-ray apparatus, a gas bubble was detected within the puparium of Glossina morsitans. The bubble appeared between 6 and 15 h after pupariation and was associated with one of the longitudinal tracheal trunks of the third instar larva. The bubble grew and achieved maximum size approximately 96 h after pupariation. It then disappeared at the time of eversion of the pupal appendages. There was a close correlation between bubble size and the weight of water lost since the time of pupariation. At the time of eversion of the pupal appendages the gas bubble apparently passed out through the longitudinal tracheal trunk and posterior spiracle to occupy the space between larval (puparial) and pupal cuticle. It is suggested that the bubble plays a vital role in the separation of these cuticular layers and that to this end water loss from the puparium is essential.     

Water loss and respiration of Lucilia cuprina during development within the puparium

The rate of loss of water and the rate of uptake of oxygen were measured continuously throughout the development of Lucilia cuprina within the puparium. Changes in these parameters were correlated with changes observed in morphology of cuticles and respiratory structures during development.In development at 26°C, there is, at 20–22 hr after puparium formation a major loss of water by mechanical expulsion of moulting fluid chiefly through the posterior larval spiracles after the severing of the posterior larval tracheae. This loss of water is essential to survival and is followed by an extremely low rate of water loss attributed to slow diffusion of water through the resulting air gap between the pupal cuticle and the puparium. There is an increase in oxygen consumption during the pupal movements associated with the casting of the larval tracheae followed by a sharp reduction in oxygen consumption until the pupal horns are everted a short time later. This combination of physiological events enables development to proceed over a wide range of conditions in the puparial environment.     

Differentially expressed genes in metamorphosis and after juvenile hormone application in the pupa of Galleria.

In Galleria mellonella, the pupal-adult transformation of epidermal cells is initiated at day 1 after pupal ecdysis by downregulation of pupal syntheses and loss of juvenile hormone (JH) sensitivity, indicating the change from pupal to adult commitment. To trace regulatory events as close as possible to the early steps of this process, we have analyzed, by differential display, changes in epidermal mRNA populations during the first day after pupal ecdysis in normal development as well as after JH injection. We isolated and cloned 20 cDNA 3'-fragments that are differentially expressed with regard to their developmental profile either in normal development or after injection of JH. Four clones could be verified by Northern blot hybridization.Screening of corresponding cDNA libraries with digoxigenin-labeled anti-sense mRNA probes yielded two full-length cDNA clones (9/27 and 23/86). Both of them represent genes that could be involved in the regulatory events during initiation of pupal metamorphosis or in the action of JH, respectively. The 9/27 mRNA is inducible by JH. It contains, in the 3' untranslated region, a consensus sequence for deadenylation and specific degradation. The corresponding protein possesses two PKC phosphorylation sites and is with high probability a nuclear protein. The 23/86 clone represents polyubiquitin, differentially regulated in normal development and after JH application.     

Role of Tau,a microtubule associated protein,in Drosophila photoreceptor morphogenesis

Cell polarity genes have important functions in photoreceptor morphogenesis. Based on recent discovery of stabilized microtubule cytoskeleton in developing photoreceptors and its role in photoreceptor cell polarity, microtubule associated proteins might have important roles in controlling cell polarity proteins' localizations in developing photoreceptors. Here, Tau, a microtubule associated protein, was analyzed to find its potential role in photoreceptor cell polarity. Tau colocalizes with acetylated/stabilized microtubules in developing pupal photoreceptors. Although it is known that tau mutant photoreceptor has no defects in early eye differentiation and development, it shows dramatic disruptions of cell polarity proteins, adherens junctions, and the stable microtubules in developing pupal photoreceptors. This role of Tau in cell polarity proteins' localization in photoreceptor cells during the photoreceptor morphogenesis was further supported by Tau's overexpression studies. Tau overexpression caused dramatic expansions of apical membrane domains where the polarity proteins localize in the developing pupal photoreceptors. It is also found that Tau's role in photoreceptor cell polarity depends on Par‐1 kinase. Furthermore, a strong genetic interaction between tau and crumbs was found. It is found that Tau has a crucial role in cell polarity protein localization during pupal photoreceptor morphogenesis stage, but not in early eye development including eye cell differentiation.     

Developmental expression and hormonal regulation of different isoforms of the transcription factor E75 in the tobacco hornworm Manduca sexta

E75A and E75B, isoforms of the E75 orphan nuclear receptor, are sequentially up-regulated in the abdominal epidermis of the tobacco hornworm Manduca sexta by 20-hydroxyecdysone (20E) during larval and pupal molts, with E75A also increasing at pupal commitment (Zhou et al., Dev. Biol. 193, 127-138, 1998). We have now cloned E75C and show that little is expressed in the epidermis during larval life with trace amounts seen just before ecdysis. Instead, E75C is found in high amounts during the development of the adult wings as the ecdysteroid titer is rising, and this increase was prevented by juvenile hormone (JH) that prevented adult development. By contrast, E75D is expressed transiently during the larval and pupal molts as the ecdysteroid titer begins to decline and again just before ecdysis, but in the developing adult wings is expressed on the rise of 20E. Removal of the source of JH had little effect on either E75C or E75D mRNA expression during the larval and pupal molts. At the time of pupal commitment, in vitro experiments show that 20E up-regulates E75D and JH prevents this increase. Neither E75A nor E75D mRNA was up-regulated by JH alone. Thus, E75C is primarily involved in adult differentiation whereas E75D has roles both during the molt and pupal commitment.     

Water Level Flux in Household Containers in Vietnam - A Key Determinant of Aedes aegypti Population Dynamics

We examined changes in the abundance of immature Aedes aegypti at the household and water storage container level during the dry-season (June-July, 2008) in Tri Nguyen village, central Vietnam. We conducted quantitative immature mosquito surveys of 171 containers in the same 41 households, with replacement of samples, every two days during a 29-day period. We developed multi-level mixed effects regression models to investigate container and household variability in pupal abundance. The percentage of houses that were positive for I/II instars, III/IV instars and pupae during any one survey ranged from 19.5-43.9%, 48.8-75.6% and 17.1-53.7%, respectively. The mean numbers of Ae. aegypti pupae per house ranged between 1.9-12.6 over the study period. Estimates of absolute pupal abundance were highly variable over the 29-day period despite relatively stable weather conditions. Most variability in pupal abundance occurred at the container rather than the household level. A key determinant of Ae. aegypti production was the frequent filling of the containers with water, which caused asynchronous hatching of Ae. aegypti eggs and development of cohorts of immatures. We calculated the probability of the water volume of a large container (>500L) increasing or decreasing by ≥20% to be 0.05 and 0.07 per day, respectively, and for small containers (<500L) to be 0.11 and 0.13 per day, respectively. These human water-management behaviors are important determinants of Ae. aegypti production during the dry season. This has implications for choosing a suitable Wolbachia strain for release as it appears that prolonged egg desiccation does not occur in this village.     

Relation between cell number and pupal development of wing disks in Bombyx mori

The effects of JH and ecdysone on pupal differentiation of the wing disk of Bombyx mori were studied in vivo and in vitro. JH prevents pupal differentiation during larval life by stopping a particular stage of the cell cycle. Immediately after allatectomy, a cell cycle sets in without ecdysone, but afterwards wing disks obtain the competence to differentiate to the pupal type in response to ecdysterone. Disks older than 2 days after allatectomy can develop to the pupal type with an abrupt increase of mitosis in a certain concentration of ecdysterone in vitro. Once the disks have gained such competence and begun pupal development, JH no longer exhibits the effect that prevents DNA synthesis, which is enhanced by ecdysterone. It is therefore suggested that there are two phases in DNA synthesis: one which is important for achieving competence and is inhibited by JH and relatively independent of ecdysone; and another which is important for morphogenetic development and depends on ecdysone but is not inhibited by JH.     

The effect of water quality in the life cycle and in the attraction for the egg oviposition of Aedes aegypti (L.) (Diptera: Culicidae)

The present research aimed at evaluating the influence of the water quality in the life cycle and attraction of Aedes aegypti (L.) females to oviposit using different sources of water (raw sewage, effluent of UASB reactor, effluent of polishing lagoon, effluent of anaerobic filter, rain water and de-chlorinated water). The immature development time and survivorship were evaluated on a daily basis in two distinct feeding systems (with and without food). The quality of the water was shown to affect the egg and larval stages, but not the pupal or the adult. In the absence of food, no development was observed in rain water and de-chlorinated water. Immature development was faster in water sources from raw sewage, although with the lowest survivorship (37.3%). Free-choice tests indicated that females preferred to lay most of their eggs on water collected from the effluent of a UASB reactor, achieving the highest oviposition activity index (OAI) of 0.57. In non-choice tests, females laid larger batches of eggs in water collected from anaerobic filters (204.8 eggs), with the lowest number of eggs being laid on de-chlorinated water (37.3 eggs). It can be concluded that A. aegypti does not demonstrate any particular preference to lay eggs on clean water. This has serious implications for developing strategies to manage populations of this important vector in urban areas as it was shown to lay eggs and successfully develop on several different sources of water.     

Ecdysteroid synthesis by the ring gland of Drosophila melanogaster during late-larval,prepupal and pupal development

Radioimmunoassay has been used to determine the characteristics of ecdysteroid synthesis by ring glands and brain-ring gland preparations from late 3rd-instar larvae of Drosophila melanogaster cultured in vitro. The rate of synthesis and secretion is linear for at least 4 hr in culture. Using a 4-hr culture period, variation in the rate of ecdysteroid synthesis by brain-ring gland preparations during larval, prepupal and pupal development has been examined. The rate of synthesis and secretion is highest in late 3rd-instar larvae and decreases after puparium formation. During pupal development, at a time when the endogenous ecdysteroid titre is again increasing, the rate of ecdysteroid synthesis by brain-ring gland preparations remains low and is only 10% of that prior to puparium formation. It is, therefore, likely that the ring gland is not a major source of ecdysteroids during this period.     

A model of pupal water loss in Glossina

The results of a long-established investigation into pupal transpiration are used as a rudimentary data set. These data are then generalised to all temperatures and humidities by invoking the property of multiplicative separability, as well as by converting established relationships in terms of constant humidity at fixed temperature, to alternatives in terms of a calculated water loss. In this way a formulation which is a series of very simple, first order, ordinary differential equations is devised. The model is extended to include a variety of Glossina species using their relative surface areas, their relative pupal and puparial loss rates and their different 4th instar excretions. The resulting computational model calculates total, pupal water loss, consequent mortality and emergence. Remaining fat reserves are a more tenuous result.The model suggests that, while conventional wisdom is often correct in dismissing variability in transpiration-related pupal mortality as insignificant, the effects of transpiration can be profound under adverse conditions and for some species, in general. The model demonstrates how two gender effects, the more significant one at the drier extremes of tsetse fly habitat, might arise. The agreement between calculated and measured critical water losses suggests very little difference in the behaviour of the different species.     

Light and electron microscopic studies on the neurosecretory control of diapause incidence in Pieris rapae crucivora

The incidence of diapause was shown to be determined humorally during the larval-pupal ecdysis by means of brain extirpation experiments.On the basis of this observation, light and electron microscopic changes in the neurosecretory type II cells in the pars intercerebralis-corpus cardiacum system during pharate pupal and early pupal stages were examined in insects reared under long day-length (non-diapause individuals) and in insects reared under short day-length (diapause individuals). In the diapause individuals, neurosecretory granules in NS-II cells increased during the pupal instar and large aggregates of granules packed the cytoplasm. Thereafter, inclusion bodies showing cytoplasmic breakdown of the granules appeared.In the non-diapause individuals, on the contrary, electron micrographs suggesting the release of neurosecretory material from axon terminals were obtained just after the pupal ecdysis. There were very few granules, with many Golgi bodies and much rough ER 8 to 12 hr after the ecdysis.It is concluded that adult development is determined by the release of neurosecretory material from the axon terminals of NS-II cells at the larval-pupal ecdysis. If release does not occur, the pupae enter diapause. It is also thought that differences in day-length during the larval stages influence the activities of NS-II cells before pupation.     

Genetic and phenotypic variation in diet breadth in a generalist herbivore

Summary The genetic and plastic components of polyphagy were investigated in a population ofLymantria dispar, the gypsy moth. A simple genetic experiment assessed the expression of (1) genetic variability in life history traits within each of four environments, (2) genetic variability in diet breadth, expressed as a change in the ranks of family performance across hosts, and (3) homeostasis (equivalent performance by a family across hosts) versus phenotypic plasticity (variable performance by a family across hosts). Sibs from each of 14 families, randomly selected from a single population, were reared on four diets: two natural hosts — chestnut and red oak, and two synthetic hosts — a standard laboratory diet and a low-protein version of this diet. Average population performance, measured in terms of development time and pupal weight, was better on standard laboratory diet than on low-protein diet, and was equal on chestnut and red oak for pupal weight, but better on chestnut oak for development time. Average population performance provided no information about the genetic component of host use ability. The gypsy moth expressed genetic variation in development time within each host environment and in pupal weight within natural host environments. Phenotypic plasticity was expressed by a significant number of families in development time and pupal weight across synthetic hosts and, to a lesser extent, across natural hosts. It was only across natural hosts that genetic variation in diet breadth was expressed, and this was confined to females. Genetic variability in diet breadth may be maintained in this species as a consequence of the unpredictability of its food sources.     

栎黄枯叶蛾生长发育与几种寄主内含物的关系

【目的】探讨饲喂不同寄主植物对栎黄枯叶蛾Trabala vishnou gigantina Yang生长发育及繁殖的影响,分析寄主植物内含物与其生长发育及繁殖的关系,为研究不同寄主植物对栎黄枯叶蛾种群动态的影响提供理论依据。【方法】在室内条件下,利用沙棘Hippophae rhamnoides、山杏Armeniaca sibirica、山杨Populus davidiana、旱柳Salix matsudana饲养该虫,观察幼虫发育历期、蛹历期、蛹重及产卵量等指标,同时测定寄主叶片中可溶性糖、可溶性蛋白质、水分、单宁及黄酮的含量,并进行线性回归分析。【结果】不同寄主植物饲喂栎黄枯叶蛾幼虫的发育历期、蛹历期、蛹重及产卵量均存在显著差异。取食沙棘的幼虫历期最短,仅为(74.50?1.76)d,而取食旱柳历期最长,达(106.00?1.51)d。取食山杨的蛹历期最长,为(44.00?1.41)d,最短的是取食旱柳的,仅为(32.70?3.13)d。饲喂沙棘和山杏的雌、雄蛹重和成虫产卵量显著高于饲喂山杨和旱柳的。对不同寄主的营养物质、次生物质和水分含量测定结果表明:不同寄主间营养物质、次生物质与水分的含量差异显著。回归相关分析表明:幼虫发育历期与寄主植物可溶性蛋白质含量呈显著负相关;雌蛹重和产卵量与寄主植物可溶性糖含量呈显著负相关;雌、雄蛹重与寄主植物含水量呈显著正相关;蛹历期与单宁含量呈显著正相关,但黄酮含量的高低与栎黄枯叶蛾生长发育等指标无相关性。【结论】沙棘是栎黄枯叶蛾生长发育及繁殖的最适寄主植物;并且寄主植物中可溶性糖、单宁的含量低、含水量和可溶性蛋白质含量高有利于栎黄枯叶蛾的生长发育和繁殖。     

A reverse temperature‐size rule associated with a negative relationship between larval development time and pupal weight in a tropical population of Ostrinia furnacalis

Empirical studies demonstrate a negative relationship between body size and temperature [i.e. the temperature‐size rule (TSR)] and a positive relationship between body size and development time (a trade‐off). However, many exceptions are also reported. The present study tests the two relationships in a tropical population of the Asian corn borer Ostrinia furnacalis under a wide range of temperatures from 18 to 31 °C. The results show that larval time is decreased significantly and pupal weight is increased significantly with an increasing rearing temperature, exhibiting the reverse‐TSR. Growth rate is positively correlated with temperature. The relationships between body size and development time among individuals show a negative slope (i.e. pupal weight tends to decrease with increases in development time). Females are significantly larger than males at all temperatures, showing a female‐biased sex size dimorphism. It is also found that development time and pupal weight vary greatly among individuals, particularly at lower temperatures. The results of the present study demonstrate a reverse‐TSR associated with a negative relationship between larval development time and pupal weight in a tropical population of O. furnacalis.     

家蚕组织蛋白酶基因家族的鉴定及表达特征分析

家蚕是鳞翅目完全变态昆虫,在其变态过程中伴随着巨大的形态变化,包括旧组织的解离和新组织的形成,在这过程中有多种组织蛋白酶参与。组织蛋白酶是一类细胞内蛋白酶,广泛存在于各个物种中,包括组织蛋白酶B、H、L等几个亚家族。对家蚕组织蛋白酶的研究将有利于阐明家蚕变态发育的详细过程。通过对家蚕基因组数据库进行筛选,共在家蚕中鉴定到13种组织蛋白酶,并对这13种组织蛋白酶的基本信息和表达模式进行了分析。另外,利用家蚕基因芯片数据和荧光定量PCR分析,鉴定编号为BGIBMGA004622的基因为卵巢特异表达的组织蛋白酶L亚家族基因。该基因全长1 209 bp,编码402个氨基酸。经过序列分析,该酶与其他物种的组织蛋白酶L具有较高的同源性,其活性位点高度保守,且与鳞翅目的组织蛋白酶L在进化上聚为一支。同时,对该基因进行克隆并原核表达,结果显示重组蛋白以包涵体的形式表达。定量PCR结果显示,该酶在蛹发育初期表达量逐渐升高,至蛹3 d达到最高值,推测其可能参与卵巢与卵母细胞的发育过程。     

Endocrine Mechanisms Regulating Post-Diapause Development in the Cabbage Armyworm,Mamestra brassicae

Diapause, a programmed developmental arrest at a specific stage, is common in insects and is regulated by hormones. It is well established that in pupal diapause, cessation of ecdysteroid secretion from the prothoracic glands (PGs) after pupal ecdysis leads to diapause initiation, while resumption of its secretion induces post-diapause development. However, what regulates the activity of the glands is poorly understood, especially for the glands of diapause-terminated pupae. In the present study, we investigate the mechanisms by which post-diapause development is regulated in the cabbage armyworm Mamestra brassicae. We demonstrate that the brain is necessary for the initiation of post-diapause development and that the factor in the brain responsible for the activation of the PGs is the prothoracicotropic hormone (PTTH). Further, through measuring the hemolymph PTTH titers by time-resolved fluoroimmunoassay, we show that PTTH is actually released into the hemolymph prior to the activation of the PGs. Although its peak titer is much lower than expected, this low concentration of PTTH is most likely still effective to activate the PGs of post-diapause pupae, because the responsiveness to PTTH of the glands at this stage is very high compared to that of nondiapause pupal PGs. These results strongly suggest that in M. brassicae, PTTH serves as a trigger to initiate pupa-adult development after diapause termination by stimulating the PGs to secrete ecdysteroid.     

Northward invasion of the parasitic deer ked ( Lipoptena cervi), is there geographical variation in pupal size and development duration?

The deer ked (Lipoptena cervi) is a common ectoparasite of cervids. During the last decades the species has rapidly invaded in northern Europe, especially in Finland, towards the north and increased its prevalence on the moose population. Consequently, during this rapid invasion the deer ked has faced more severe climatic conditions. We studied whether pupal size (measured as pupal weight) and pupal development duration of the deer ked varies along historical invasion zones and temperature zones towards north in Finland. Moreover, we explored possible size- and gender-dependent variation in pupal development duration. We divided wild-collected pupae in respect to their origin in two ways: (1) temperature zones (from south-west to colder north-east) and (2) invasion history (from early to late establishment). We reared pupae in the controlled laboratory conditions in identical temperature and light conditions. Pupal size decreased towards north and the smaller pupae developed faster. However, the results do not show differences in pupal size or developmental characteristics between the invasion zones. This supports the idea of rapid developmental plasticity of the deer ked and that not the invasion history but the current temperature regime determines the life history of the deer ked when invading towards a colder environment.     

Patterning during pupal commitment of the epidermis in the butterfly, Precis coenia: the role of intercellular communication

The commitment of cells to pupal development in the larvae of holometabolous insects can be prevented by treatment with juvenile hormone (JH) or a JH mimic during a critical period early in the last larval instar. By treating larvae of different ages with a JH mimic, pupal commitment of the epidermis of the butterfly, Precis coenia, was found to occur in a strict temporal and spatial progression, which was serially homologous and occurred independently in each segment. The mechanism underlying this sequence of pupal commitment was examined by cauterizing regions of the epidermis to observe the effects of local ablation on the pattern of pupal commitment revealed by treatment with the JH mimic. Cautery of the segmental site of origin of pupal commitment, the dorsal midline, suppressed pupal commitment in the rest of the operated segment, indicating that the midline has a special effect on commitment of the rest of the segment. Cautery off the midline produced asymmetries in the pattern of pupal commitment; when placed close to the midline, such cauteries prevented pupal commitment in the region "downstream" of the cautery, suggesting that a signal (diffusible or transducible) emanates from the midline. Finally, cautery of a circle around the midline inhibited pupal commitment only outside the circle, showing that cautery could act as a barrier to the passage of a signal coming from the midline. These results suggest that inductive as well as hormonal signals are involved in the regulation of pupal commitment in the epidermis of the lepidopteran, P. coenia.