Final Larval Stage Research Articles

Morphometric relationships and seasonal variation in size, weight, and a condition index of post-settlement stages of the Caribbean spiny lobster.

Spiny lobsters have a protracted pelagic, oceanic larval phase. The final larval stage metamorphoses into a non-feeding postlarva (puerulus) that actively swims towards the coast to settle in shallow habitats and does not resume feeding until after the molt into the first-stage juvenile. Therefore, the body dimensions and nutritional condition of both settled pueruli and first juveniles are likely to vary over time, potentially playing a crucial role in the recruitment to the benthic population. We compared carapace length (CL), height (CH), and width (CW); total length (TL), and body weight (W) between pueruli and first juveniles of the Caribbean spiny lobster, Panulirus argus, as well as morphometric relationships between both developmental stages. Except for CL, all other dimensions were larger in first juveniles, but more markedly CH and W. The slopes of the CH vs CL, CW vs CL, and W vs CL regressions differed significantly between stages, and all log-transformed relationships showed isometry in both stages, except for the CH vs CL relationship, which showed positive allometry. These results reflect a morphological change from the flatter, more streamlined body of the puerulus, to the heavier, more cylindrical body of the juvenile. We also analyzed seasonal variations in CL, W, the W/CL index (a morphometric condition index), and a modified W/CL index (i.e. after controlling for a significant effect of CL) of both stages using individuals monthly collected over 12 consecutive seasons (Autumn 2010–Summer 2013). In both stages, all three variables exhibited significant seasonal variation. For pueruli, the modified W/CL index differed from average in only two seasons, winter 2011 (higher) and summer 2013 (lower), but showed great within-season variation (larger coefficients of variation, CV), potentially reflecting variability in nutritional condition of larvae prior to metamorphosis and in the distances swum by individual pueruli to the settlement habitats. For first juveniles, the modified W/CL index was higher than average in winter and spring 2011, and lower in autumn 2011 and winter 2012, but showed less within season variation (smaller CVs), suggesting a combination of carry-over effects of puerulus condition and effects of local conditions (e.g., food availability and predation risk). These findings warrant further investigation into factors potentially decoupling settlement from recruitment processes.

Evidence for a Prepupal Diapause in the Mountain Pine Beetle (Dendroctonus ponderosae).

Dormancy strategies, including diapause and quiescence, enable insects to evade adverse conditions and ensure seasonally appropriate life stages. A mechanistic understanding of a species' dormancy is necessary to predict population response in a changing climate. Climate change is influencing distribution patterns and population success of many species, including Dendroctonus ponderosae Hopkins (Coleoptera: Curculionidae: Scolytinae), the most important mortality agent of pines in western North America. Diapause is considered absent in D. ponderosae, and quiescence in the final larval stage prior to pupation (i.e., prepupal) is considered the main dormancy strategy. We evaluated if a facultative diapause in the prepupal stage, rather than a pupation threshold ~15°C (i.e., quiescence), could describe pupation patterns in two latitudinally separated D. ponderosae populations in the western United States. We hypothesized that if pupation occurs at lower temperatures than previously described, and if significant prepupal developmental delays occur, diapause is a likely physiological mechanism. Although there was considerable variation within and between populations, pupation occurred below the previously established threshold suggesting a prepupal facultative diapause that is induced when late instars experience cool temperatures. Individuals that pupated at temperatures below 15°C also had developmental delays, relative to development at warmer temperatures, consistent with diapause development. Pupation patterns differed between populations wherein diapause was induced at cooler temperatures and diapause development was shorter in southern compared with northern D. ponderosae. Recognition of a facultative diapause that varies among and between populations is critical for making predictions about future population response and range expansion in a changing climate.

The influence of oceanographic conditions and larval behaviour on settlement success—the European sea bass Dicentrarchus labrax (L.)

Abstract The European Seabass (Dicentrarchus labrax) is a slow-growing late maturing fish. The northern stock has been declining since 2010 and is thought to be caused by a combination of fishing and weak year classes. Large inter-annual variation in settlement has been observed, so a better understanding of the mechanisms driving settlement success will aid interpretation of the variation between years, and help to improve the stock assessment models and management strategies. In this study, an individual-based model (IBM) was developed to investigate the factors affecting sea bass settlement on nursery grounds of the northern sea bass stock. The IBM was coupled with hydrodynamics to track particles, whereas egg and larval development, and vertical migration behaviour are fully incorporated. The IBM successfully predicted inter-annual differences in settlement regardless of larval behaviour. The highest settlement success was predicted with neutrally buoyant eggs, hatchlings, and larval stages, in combination with tidal migration at the final larval stage. Dispersal was driven mainly by the influence of wind on residual currents and water temperature, with warmer temperatures reducing the duration of the pelagic phase and stronger current increasing the potential to drift further. Eggs spawned in the central western English Channel settled in both England and France, with movement from the central to the eastern English Channel occurring only in warm years. Larval duration was driven by water temperature and showed an increase in duration from the southwest to northeast areas of the northern stock. The results are discussed in the context of sea bass management and conservation strategies.

RNAi-mediated knockdown of the voltage gated sodium ion channel TcNav causes mortality in Tribolium castaneum.

The voltage-gated sodium ion channel (VGSC) belongs to the largest superfamily of ion channels. Since VGSCs play key roles in physiological processes they are major targets for effective insecticides. RNA interference (RNAi) is widely used to analyse gene function, but recently, it has shown potential to contribute to novel strategies for selectively controlling agricultural insect pests. The current study evaluates the delivery of dsRNA targeted to the sodium ion channel paralytic A (TcNav) gene in Tribolium castaneum as a viable means of controlling this insect pest. Delivery of TcNav dsRNA caused severe developmental arrest with larval mortalities up to 73% post injection of dsRNA. Injected larvae showed significant (p < 0.05) knockdown in gene expression between 30–60%. Expression was also significantly (p < 0.05) reduced in pupae following injection causing 30% and 42% knockdown for early and late pupal stages, respectively. Oral delivery of dsRNA caused dose-dependant mortalities of between 19 and 51.34%; this was accompanied by significant (p < 0.05) knockdown in gene expression following 3 days of continuous feeding. The majority of larvae injected with, or fed, dsRNA died during the final larval stage prior to pupation. This work provides evidence of a viable RNAi-based strategy for insect control.

Development and functional morphology of the mouthparts and foregut in larvae and post-larvae of Macrobrachium jelskii (Decapoda: Palaemonidae)

The morphology of the mouthparts and foregut of the larvae and post-larvae of Macrobrachium jelskii was investigated to determine their functional roles in feeding, in order to understand the larval feeding behaviour and the changes that occur during its development. The mouthparts and foregut of the zoea I and II are morphologically similar, rudimentary and non-functional in feeding. Only in the final larval stage, zoea III, do the external mouthparts and foregut become structurally more complex and thus likely to play a potential role in feeding. Two behavioral trials (point of no return, point of reserve saturation) evaluated the resistance to starvation in zoea I, II, and III. The results indicate that they have sufficient nutritional reserves to permit them to complete metamorphosis without feeding. Overall, our results suggest that the zoea I and II of Macrobrachium jelskii engage in obligate lecithotrophy and zoea III in facultative lecithotrophy.

Ectopic expression of ecdysone oxidase impairs tissue degeneration in Bombyx mori.

Metamorphosis in insects includes a series of programmed tissue histolysis and remolding processes that are controlled by two major classes of hormones, juvenile hormones and ecdysteroids. Precise pulses of ecdysteroids (the most active ecdysteroid is 20-hydroxyecdysone, 20E), are regulated by both biosynthesis and metabolism. In this study, we show that ecdysone oxidase (EO), a 20E inactivation enzyme, expresses predominantly in the midgut during the early pupal stage in the lepidopteran model insect, Bombyx mori. Depletion of BmEO using the transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases) system extended the duration of the final instar larval stage. Ubiquitous transgenic overexpression of BmEO using the Gal4/UAS system induced lethality during the larval-pupal transition. When BmEO was specifically overexpressed in the middle silk gland (MSG), degeneration of MSG at the onset of metamorphosis was blocked. Transmission electron microscope and LysoTracker analyses showed that the autophagy pathway in MSG is inhibited by BmEO ectopic expression. Furthermore, RNA-seq analysis revealed that the genes involved in autophagic cell death and the mTOR signal pathway are affected by overexpression of BmEO. Taken together, BmEO functional studies reported here provide insights into ecdysone regulation of tissue degeneration during metamorphosis.

FRESHWATER UNIONID BIVALVE SHELLS AS SUBSTRATA FOR TRICHOPTERA ATTACHMENT

ABSTRACT A variety of competitive advantages accompany colonization of hard substrates in aquatic settings, and as a result, many organisms adopt this mode of life. Similarly, adverse effects may occur to organisms that have themselves been encrusted. Many marine basibiont faunas (organisms that provide a substrate for populations of attached epibiontic organisms), including bivalve mollusks, are known to employ a range of chemical and physical deterrents to minimize the incidence of epibiotic attachment, with its attendant detrimental impacts. Freshwater basibiont and epibiont associations, however, are poorly documented. We collected an assemblage of dead, empty freshwater mussel (unionid bivalve) shells from the Saint John River, New Brunswick, Canada. Some unionid specimens exhibited colonization by attached Trichoptera (caddisfly) cases, in either their final instar larval or pupal developmental stages. The trichopteran assemblage, identified from case morphology, included abundant Neophylax sp. toge...

CYP18A1 regulates tissue-specific steroid hormone inactivation in Bombyx mori

Insect development and metamorphosis are regulated by two major hormones, juvenile hormone and ecdysteroids. Despite being the key regulator of insect developmental transitions, the metabolic pathway of the primary steroid hormone, 20-hydroxyecdysone (20E), especially its inactivation pathway, is still not completely elucidated. A cytochrome P450 enzyme, CYP18A1, has been shown to play key roles in insect steroid hormone inactivation through 26-hydroxylation. Here, we identified two CYP18 (BmCYP18A1 and BmCYP18B1) orthologs in the lepidopteran model insect, Bombyx mori. Interestingly, BmCYP18A1 gene is predominantly expressed in the middle silk gland (MSG) while BmCYP18B1 expresses ubiquitously in B. mori. BmCYP18A1 is induced by 20E in vitro, suggesting its role in 20E metabolism. Using the binary Gal4/UAS transgenic system, we ectopically overexpressed BmCYP18A1 in a MSG-specific manner with a Sericin1-Gal4 (Ser-Gal4) driver or in a ubiquitous manner with an Actin3-Gal4 (A3-Gal4) driver. Ectopic overexpression of BmCYP18A1 in MSG or in all tissues resulted in developmental arrestment of transgenic animals during the final instar larval stage. The 20E titers in the transgenic animals expressing BmCYP18A1 were lower compared to the levels in the control animals. Although the biological significance of MSG-specific expression of BmCYP18A1 is unclear, our results provide the first evidence that BmCYP18A1, which is conserved in most arthropods, is involved in a tissue-specific steroid hormone inactivation in B. mori.

Suscetibilidade de Larvas de Simulídeos ao Larvicida Temephos em Caarapó, MS

Blackfly control in Brazil is necessary to avoid nuisance to rural or urban communities and economic losses in agricultural or tourist activity areas. The susceptibility of Simulium spp. larvae was performed to the organophosphate temephos (Abate 500 CE) at the Caarapo stream, in the city of Caarapo in the state of Mato Grosso do Sul. The resistance to the larvicide was indicated by the rapid biochemical test of esterase activity carried out with extracts of larvae, and confirmed by the results of the field bioassay with larvae in troughs in the stream bed. For the field bioassay final stage larvae were exposed to a Temephos Operational Concentration of 0.5 ppm ai/10 minutes and a Diagnostic Concentration of 2.5 ppm ai/10 minutes and low mean mortality was obtained (5.7% and 44.6 % respectively), confirming the resistance and indicating the need for using other products for blackfly control in this area.

In situ swimming and orientation behavior of spiny lobster (Panulirus argus) postlarvae

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 504:207-219 (2014) - DOI: https://doi.org/10.3354/meps10748 In situ swimming and orientation behavior of spiny lobster (Panulirus argus) postlarvae Andrew S. Kough*, Claire B. Paris, Erica Staaterman Rosenstiel School of Marine and Atmospheric Science, University of Miami, Miami, Florida 33149, USA *Corresponding author: andrew.kough@gmail.com ABSTRACT: Larval dispersal is common in benthic marine organisms, but the environmental cues guiding the final larval stages to settlement habitat remain largely unknown. Caribbean spiny lobster Panulirus argus spend 5 to 12 mo at sea as pelagic phyllosome larvae before transitioning into rapidly swimming and non-feeding puerulus postlarvae. Spiny lobster postlarvae are specialized to seek settlement habitat, making them an ideal model for studying orientation behavior during the transition to benthic environments. We observed the swimming orientation of P. argus in the open ocean during all hours of the day and across tidal phases using a circular behavioral arena deployed at sea. Contrary to laboratory observations of only nocturnal activity, swimming and orientation occurred during both the day and the night in open waters ca. 6 to 8 km offshore in the Florida Straits. Individual postlarvae swam directionally with similar accuracy in all hours. The mean orientation direction was significantly different with tidal phase. Postlarvae oriented to the southeast (163.5°) during flood tide, and to the southwest during ebb flow (225.1°), placing them on a shoreward trajectory. Postlarval behavior was circatidal, and during ebb tide, innate swimming behavior beneficial to finding settlement habitat was triggered. Postlarvae also oriented relative to the wind, potentially using the sea breeze to orient towards the coast. A change in behavior tied to ubiquitous coastal signals is a strategy likely used by other pelagic larvae searching for a benthic home. KEY WORDS: Spiny lobster · Orientation · Postlarva · Nocturnal · Tidal · Dispersal · Settlement Full text in pdf format PreviousNextCite this article as: Kough AS, Paris CB, Staaterman E (2014) In situ swimming and orientation behavior of spiny lobster (Panulirus argus) postlarvae. Mar Ecol Prog Ser 504:207-219. https://doi.org/10.3354/meps10748 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 504. Online publication date: May 14, 2014 Print ISSN: 0171-8630; Online ISSN: 1616-1599 Copyright © 2014 Inter-Research.

Aging increases resistance to bacterial infection in the tobacco hornworm, Manduca sexta (1100.4)

Manduca sexta larvae (tobacco hornworm) early in their fifth and final larval stage have been reported to have more robust immune responses to Photorabdus luminscens compared to animals later in that stage. We hypothesized that the robust immune response in early stage caterpillars would result in varying hemolymph bacterial loads and survival to other bacterial infections. Early stage M. sexta injected with non‐pathogenic E. coli had 25‐fold higher bacterial loads in hemolymph than those injected later. In addition, early stage M. sexta injected with pathogenic Stenotrophomonas maltohphilia died twice as fast as those injected later. These data demonstrate an ineffective immune response in early stage caterpillars. We also hypothesized that the robust immune response in early stage caterpillars would affect other aspects of physiology, such as development and growth. Early stage M. sexta injected with E. coli or S. maltophilia exhibited developmental and growth rates twice as slow as those injected later. Sickness‐induced anorexia occurs in many taxa and may be the cause of the growth rate and developmental delays in infected caterpillars. While it is clear that development affects immunity, it is unclear why the age‐immunity relationship varies with different bacteria. Because of the importance of insects as pests and pollinators, understanding how immunity varies throughout development is critical.

Effects of various sodium adsorption ratio (SAR) mediums on larval performance of the freshwater prawnMacrobrachium rosenbergii(de Man)

AbstractThe sodium adsorption ratio (SAR) is used to describethe ratio of sodium to calcium and magnesium inwater that is very important in both aquacultureand agriculture. SAR treatments including of 20, 30,40, 50 and 60 were adjusted in 10-L containers. Ini-tial larval density was fixed at 100 larvae L 1 andduring the culture period, various larval qualityparameters were evaluated. Our findings illustratedthat at initial stages of larval development, differentSAR treatments affected the larval condition index(LCI), but no influences on larval stage index (LSI)were apparent. Importantly, this process at the finallarval stages became reversed. In addition, survivalat SAR 30 was 12.4% greater than among any othertreatments. The highest larval resistance to stress(191 5 ppm), as measured by the 24-h LC 50 for-malin test also was observed for SAR 30, at the 11thlarval stage. Analysis indicated that calcium absorp-tion during larval developmental stages is affected bySAR levels in the environment and also showed thatthe optimal SAR treatment was 30 for Macrobrachi-um rosenbergii larviculture in inland hatchery brack-ish water. According to our findings and desired SARfor the environment and agricultural soil (<15),effluent of M. rosenbergii inland hatcheries must betreated at least two times by dilution and addingcalcium and magnesium salts.Keywords: Macrobrachium rosenbergii, sodiumadsorption ratio, larval stage index, larval condi-tion index, larvicultureIntroductionThe freshwater prawn Macrobrachium rosenbergii isa classic example of an aquatic species that hasbecome widely cultured because of its suitabilityfor commercialization. By 2009, global productionhad risen to 229 419 t, more than 80 times asmuch as in 1980. Presently, commercial prawnproduction continues to climb in nearly all coun-tries currently engaged in its culture (FAO Fisher-ies & Aquaculture Department 2011; New & Nair2012). This is due to a number of factors relatedto its ease of production, including the use oforganic fertilization strategies instead of expensivecomplete diets, general tolerance to low waterquality conditions and overall ease in nurseryproduction (Cavalli, Lavens & Sorgeloos 1999).However, some critical challenges do remain, espe-cially in the larviculture phase, related to both sur-vival and overall larval quality (Bart & Yen 2003).The larval phase of M. rosenbergii is fairlyunique among crustaceans. The 11-stage processis completed entirely in brackish water, whereasits other life cycle stages occur principally in fresh-water. Damrongphol, Jaroensastraraks and Pool-sanguan (2001) indicated that ionic requirementsof newly hatched M. rosenbergii larvae differedfrom that of developing embryos. Although maxi-mal growth of an aquatic organism would beassumed to occur in isosmotic media, research hasdemonstrated that this principle is not true for thegiant freshwater prawn. It was found (Singh1980) that M. rosenbergii actually had maximumgrowth occur in environmental conditions lower

Integrated analysis of cytochrome P450 gene superfamily in the red flour beetle, Tribolium castaneum

BackgroundThe functional and evolutionary diversification of insect cytochrome P450s (CYPs) shaped the success of insects. CYPs constitute one of the largest and oldest gene superfamilies that are found in virtually all aerobic organisms. Because of the availability of whole genome sequence and well functioning RNA interference (RNAi), the red flour beetle, Tribolium castaneum serves as an ideal insect model for conducting functional genomics studies. Although several T. castaneum CYPs had been functionally investigated in our previous studies, the roles of the majority of CYPs remain largely unknown. Here, we comprehensively analyzed the phylogenetic relationship of all T. castaneum CYPs with genes in other insect species, investigated the CYP6BQ gene cluster organization, function and evolution, as well as examined the mitochondrial CYPs gene expression patterns and intron-exon organization.ResultsA total 143 CYPs were identified and classified into 26 families and 59 subfamilies. The phylogenetic trees of CYPs among insects across taxa provided evolutionary insight for the genetic distance and function. The percentage of singleton (33.3%) in T. castaneum CYPs is much less than those in Drosophila melanogaster (52.5%) and Bombyx mori (51.2%). Most members in the largest CYP6BQ gene cluster may make contribution to deltamethrin resistance in QTC279 strain. T. castaneum genome encodes nine mitochondrial CYPs, among them CYP12H1 is only expressed in the final instar larval stage. The intron-exon organizations of these mitochondrial CYPs are highly diverse.ConclusionOur studies provide a platform to understand the evolution and functions of T. castaneum CYP gene superfamily which will help reveal the strategies employed by insects to cope with their environment.

Presynaptic PI3K activity triggers the formation of glutamate receptors at neuromuscular terminals of Drosophila

Synapse transmission depends on the precise structural and functional assembly between pre- and postsynaptic elements. This tightly regulated interaction has been thoroughly characterised in vivo in the Drosophila glutamatergic larval neuromuscular junction (NMJ) synapse, a suitable model to explore synapse formation, dynamics and plasticity. Previous findings have demonstrated that presynaptic upregulation of phosphoinositide 3-kinase (PI3K) increases synapse number, generating new functional contacts and eliciting changes in behaviour. Here, we show that genetically driven overexpression of PI3K in the presynaptic element also leads to a correlated increase in the levels of glutamate receptor (GluRII) subunits and the number of postsynaptic densities (PSDs), without altering GluRII formation and assembly dynamics. In addition to GluRIIs, presynaptic PI3K activity also modifies the expression of the postsynaptic protein Discs large (Dlg). Remarkably, PI3K specifically overexpressed in the final larval stages is sufficient for the formation of NMJ synapses. No differences in the number of synapses and PSDs were detected when PI3K was selectively expressed in the postsynaptic compartment. Taken together, these results demonstrate that PI3K-dependent synaptogenesis plays an instructive role in PSD formation and growth from the presynaptic side.

Presynaptic PI3K activity triggers the formation of glutamate receptors at neuromuscular terminals of Drosophila

Synapse transmission depends on the precise structural and functional assembly between pre- and postsynaptic elements. This tightly regulated interaction has been thoroughly characterised in vivo in the Drosophila glutamatergic larval neuromuscular junction (NMJ) synapse, a suitable model to explore synapse formation, dynamics and plasticity. Previous findings have demonstrated that presynaptic upregulation of phosphoinositide 3-kinase (PI3K) increases synapse number, generating new functional contacts and eliciting changes in behaviour. Here, we show that genetically driven overexpression of PI3K in the presynaptic element also leads to a correlated increase in the levels of glutamate receptor (GluRII) subunits and the number of postsynaptic densities (PSDs), without altering GluRII formation and assembly dynamics. In addition to GluRIIs, presynaptic PI3K activity also modifies the expression of the postsynaptic protein Discs large (Dlg). Remarkably, PI3K specifically overexpressed in the final larval stages is sufficient for the formation of NMJ synapses. No differences in the number of synapses and PSDs were detected when PI3K was selectively expressed in the postsynaptic compartment. Taken together, these results demonstrate that PI3K-dependent synaptogenesis plays an instructive role in PSD formation and growth from the presynaptic side.

Biology and Life History ofBalcha indica, an Ectoparasitoid Attacking the Emerald Ash Borer,Agrilus planipennis, in North America

Balcha indica Mani and Kaul (Hymenoptera: Eupelmidae) is a solitary ectoparasitoid attacking larvae, prepupae, and pupae of the emerald ash borer, Agrilus planipennis Fairmaire (Hymenoptera: Eupelmidae). Its fecundity, oviposition rate, longevity, and development time were determined in the laboratory under standard rearing conditions (25 ± 2° C, 65 ± 10% relative humidity, and 14:10 L:D). Adults lived a mean of 59 days with a maximum of 117 days. Lifetime adult fecundity averaged 36 eggs with a maximum 94 eggs per female. The egg stage lasted for a maximum of four days with ∼ 50% eggs hatched within two days. The development time of the first instars lasted for a maximum of nine days; 50% of the first instars completed their development (i.e., molted to the next instar) within five days. Instars of the intermediate and final stage larvae (after molting of the first instars occurred) could not be distinguished until they reached the pupal stage, and 50% of those larvae pupated ∼ 62 days after adult oviposition. Under the standard rearing conditions, 50% of B. indica took ∼ 83 days to complete the life cycle (from egg to adult emergence) ranging from 47 to 129 days. These results suggest that B. indica may not have more than two generations in the mid-Atlantic and Midwest regions of United States, where normal growing seasons—with average temperature above 25° C—are normally less than six months (May–October). Because of the long life span and oviposition period of adults, however, B. indica is likely to have overlapping generations.

MAB-10/NAB acts with LIN-29/EGR to regulate terminal differentiation and the transition from larva to adult in C. elegans

In Caenorhabditis elegans, a well-defined pathway of heterochronic genes ensures the proper timing of stage-specific developmental events. During the final larval stage, an upregulation of the let-7 microRNA indirectly activates the terminal differentiation factor and central regulator of the larval-to-adult transition, LIN-29, via the downregulation of the let-7 target genes lin-41 and hbl-1. Here, we identify a new heterochronic gene, mab-10, and show that mab-10 encodes a NAB (NGFI-A-binding protein) transcriptional co-factor. MAB-10 acts with LIN-29 to control the expression of genes required to regulate a subset of differentiation events during the larval-to-adult transition, and we show that the NAB-interaction domain of LIN-29 is conserved in Kruppel-family EGR (early growth response) proteins. In mammals, EGR proteins control the differentiation of multiple cell lineages, and EGR-1 acts with NAB proteins to initiate menarche by regulating the transcription of the luteinizing hormone β subunit. Genome-wide association studies of humans and various studies of mouse recently have implicated the mammalian homologs of the C. elegans heterochronic gene lin-28 in regulating cellular differentiation and the timing of menarche. Our work suggests that human homologs of multiple C. elegans heterochronic genes might act in an evolutionarily conserved pathway to promote cellular differentiation and the onset of puberty.

Histological studies on ovary differentiation in Yemini queen honeybees, Apis mellifera jemenitica (Hymenoptera: Apidae), during post-embryonic development

The histological structure of the ovary of queens of the Yemeni honeybee race, Apis mellifera jemenitica, was studied in the third and fifth larval instars, 1-, 2- and 3-day old pupae and newly emerged imago queen bees. Queen bee larvae were fed with an intensive royal food throughout the larval stage to avoid compounding regression to worker status. Several differences were found from one developmental stage to another, as well as in the different ages of each stage. The queen bee's ovaries were observed to go through several discernable stages during metamorphosis. Large cells, from which the germ cells originate, are surrounded by tiny flat cells that form the future general shape of the ovarioles (the peritoneal cover). The queen's ovaries continue to grow and differentiate after the fifth (final) larval stage and during the three pupal stages, where the ovarioles include two areas; the germ cells and the peripheral thread. However, at emergence the germ cells are not yet mature.

Role of the Halloween genes, Spook and Phantom in ecdysteroidogenesis in the desert locust, Schistocerca gregaria

The functional characterization of the Halloween genes represented a major breakthrough in the elucidation of the ecdysteroid biosynthetic pathway. These genes encode cytochrome P450 enzymes catalyzing the final steps of ecdysteroid biosynthesis in the dipteran Drosophila melanogaster and the Lepidoptera Manduca sexta and Bombyx mori. This is the first report on the identification of two Halloween genes, spook (spo) and phantom (phm), from a hemimetabolous orthopteran insect, the desert locust Schistocerca gregaria. Using q-RT-PCR, their spatial and temporal transcript profiles were analyzed in both final larval stage and adult locusts. The circulating ecdysteroid titers in the hemolymph were measured and found to correlate well with changes in the temporal transcript profiles of spo and phm. Moreover, an RNA interference (RNAi)-based approach was employed to study knockdown effects upon silencing of both transcripts in the fifth larval stage. Circulating ecdysteroid levels were found to be significantly reduced upon dsRNA treatment.

Nutritional sensitivity of fifth instar prothoracic glands in the tobacco hornworm, Manduca sexta

Insulin-regulated growth of the prothoracic glands appears to play a critical role in timing the last larval molt, and hence metamorphosis. The present study examined insulin signaling in relation to the growth and secretory activity of prothoracic glands in the tobacco hornworm, Manduca sexta. As larvae feed during the first half of the final larval stage, the prothoracic glands grow and ecdysone secretory capacity increases. During this period of growth, we verified the presence of insulin receptor transcript in the prothoracic glands and demonstrated that the glands were responsive to insulin, as evidenced by the in vitro phosphorylation of signaling proteins in the insulin pathway such as Akt/protein kinase B and FOXO. It was predicted that starvation would reduce ecdysone secretion with concomitant changes in insulin signaling. To test this prediction, larvae were starved and changes were quantified in two nutritionally sensitive transcripts, insulin receptor and the translation inhibitor 4EBP. In glands from starved larvae, growth and ecdysone secretory capacity were reduced, and insulin receptor and 4EBP transcripts were increased. The latter changes would be expected to accompany starvation in conjunction with enhanced insulin sensitivity and reduced protein synthesis. Increased transcription of insulin receptor and 4EBP strongly suggest that nutritional deprivation reduces the secretion of endogenous insulin-like hormones. When injected with insulin, 4EBP levels in the prothoracic glands of starved larvae decreased. Thus, insulin appeared to correct starvation-induced deficits in glandular protein synthesis. However, insulin injection did not enhance ecdysone secretion. Thus, although the prothoracic glands are insulin-responsive and insulin-like hormones may promote glandular growth as larvae feed, the effects of nutritional depletion on steroidogenesis in Manduca cannot be explained solely by reduced insulin.