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Diaphorina citri serves as the primary vector for 'Candidatus Liberibacter asiaticus (CLas),' the bacterium associated with the severe Asian form of huanglongbing. CLas-positive D. citri are more fecund than their CLas-negative counterparts and require extra energy expenditure. Therefore, understanding the molecular mechanisms linking metabolism and reproduction is of particular importance. In this study, we found adipokinetic hormone (DcAKH) and its receptor (DcAKHR) were essential for increasing lipid metabolism and fecundity in response to CLas infection in D. citri. Knockdown of DcAKH and DcAKHR not only resulted in the accumulation of triacylglycerol and a decline of glycogen, but also significantly decreased fecundity and CLas titer in ovaries. Combined in vivo and in vitro experiments showed that miR-34 suppresses DcAKHR expression by binding to its 3' untranslated region, whilst overexpression of miR-34 resulted in a decline of DcAKHR expression and CLas titer in ovaries and caused defects that mimicked DcAKHR knockdown phenotypes. Additionally, knockdown of DcAKH and DcAKHR significantly reduced juvenile hormone (JH) titer and JH signaling pathway genes in fat bodies and ovaries, including the JH receptor, methoprene-tolerant (DcMet), and the transcription factor, Krüppel homolog 1 (DcKr-h1), that acts downstream of it, as well as the egg development related genes vitellogenin 1-like (DcVg-1-like), vitellogenin A1-like (DcVg-A1-like) and the vitellogenin receptor (DcVgR). As a result, CLas hijacks AKH/AKHR-miR-34-JH signaling to improve D. citri lipid metabolism and fecundity, while simultaneously increasing the replication of CLas, suggesting a mutualistic interaction between CLas and D. citri ovaries.
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Fertilidad , Hemípteros , Hormonas de Insectos , Ácido Pirrolidona Carboxílico , Transducción de Señal , Animales , Hormonas de Insectos/metabolismo , Hormonas de Insectos/genética , Femenino , Hemípteros/microbiología , Ácido Pirrolidona Carboxílico/análogos & derivados , Ácido Pirrolidona Carboxílico/metabolismo , Rhizobiaceae/fisiología , Rhizobiaceae/metabolismo , Metabolismo de los Lípidos , Ovario/microbiología , Ovario/metabolismo , MicroARNs/metabolismo , MicroARNs/genética , Hormonas Juveniles/metabolismo , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Liberibacter , OligopéptidosRESUMEN
BACKGROUND: The Endosomal Sorting Complex Required for Transport (ESCRT) is a highly conserved cellular machinery essential for many cellular functions, including transmembrane protein sorting, endosomal trafficking, and membrane scission. CHMP4B is a key component of ESCRT-III subcomplex and has been thoroughly studied in the meroistic ovaries of Drosophila melanogaster showing its relevance in maintaining this reproductive organ during the life of the fly. However, the role of the CHMP4B in the most basal panoistic ovaries remains elusive. RESULTS: Using RNAi, we examined the function of CHMP4B in the ovary of Blattella germanica in two different physiological stages: in last instar nymphs, with proliferative follicular cells, and in vitellogenic adults when follicular cells enter in polyploidy and endoreplication. In Chmp4b-depleted specimens, the actin fibers change their distribution, appearing accumulated in the basal pole of the follicular cells, resulting in an excess of actin bundles that surround the basal ovarian follicle and modifying their shape. Depletion of Chmp4b also determines an actin accumulation in follicular cell membranes, resulting in different cell morphologies and sizes. In the end, these changes disrupt the opening of intercellular spaces between the follicular cells (patency) impeding the incorporation of yolk proteins to the growing oocyte and resulting in female sterility. In addition, the nuclei of follicular cells appeared unusually elongated, suggesting an incomplete karyokinesis. CONCLUSIONS: These results proved CHMP4B essential in preserving the proper expression of cytoskeleton proteins vital for basal ovarian follicle growth and maturation and for yolk protein incorporation. Moreover, the correct distribution of actin fibers in the basal ovarian follicle emerged as a critical factor for the successful completion of ovulation and oviposition. SIGNIFICANCE: The overall results, obtained in two different proliferative stages, suggest that the requirement of CHMP4B in B. germanica follicular epithelium is not related to the proliferative stage of the tissue.
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Modern insects have inhabited the earth for hundreds of millions of years, and part of their successful adaptation lies in their many reproductive strategies. Insect reproduction is linked to a high metabolic rate that provides viable eggs in a relatively short time. In this context, an accurate interplay between the endocrine system and the nutrients synthetized and metabolized is essential to produce healthy offspring. Lipids guarantee the metabolic energy needed for egg formation and represent the main energy source consumed during embryogenesis. Lipids availability is tightly regulated by a complex network of endocrine signals primarily controlled by the central nervous system (CNS) and associated endocrine glands, the corpora allata (CA) and corpora cardiaca (CC). This endocrine axis provides hormones and neuropeptides that significatively affect tissues closely involved in successful reproduction: the fat body, which is the metabolic center supplying the lipid resources and energy demanded in egg formation, and the ovaries, where the developing oocytes recruit lipids that will be used for optimal embryogenesis. The post-genomic era and the availability of modern experimental approaches have advanced our understanding of many processes involved in lipid homeostasis; therefore, it is crucial to integrate the findings of recent years into the knowledge already acquired in the last decades. The present chapter is devoted to reviewing major recent contributions made in elucidating the impact of the CNS/CA/CC-fat body-ovary axis on lipid metabolism in the context of insect reproduction, highlighting areas of fruitful research.
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The ultrastructural study on the female reproductive system of the beetle M. brevicauda (Mordellidae) confirmed the positive correlation between the length of the sperm and the size of the female seminal receptacle (Spermatheca). The spermatheca of the species is characterized by an apical bulb-like structure where the spermathecal duct forms numerous folds filled with sperm. At this level many bacterial cells are present intermingled with the duct folds. Some are organized in large structures, such as bacteriomes, while other are single bacteriocytes. The latter are often found near the basal lamina of duct epithelium. In addition, some bacteria are visible in the cytoplasm of the duct epithelial cells. Interestingly, bacterial cells have never been observed in the duct lumen. The possible function of the bacterial cells is discussed.
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Escarabajos , Microscopía Electrónica de Transmisión , Animales , Escarabajos/ultraestructura , Femenino , Masculino , Genitales Femeninos/ultraestructura , Bacterias/ultraestructura , Espermatozoides/ultraestructura , Microscopía Electrónica de RastreoRESUMEN
Exportin 1 (XPO1) is the major karyopherin-ß nuclear receptor mediating the nuclear export of hundreds of proteins and some classes of RNA and regulates several critical processes in the cell, including cell-cycle progression, transcription and translation. Viruses have co-opted XPO1 to promote nucleocytoplasmic transport of viral proteins and RNA. Maize mosaic virus (MMV) is a plant-infecting rhabdovirus transmitted in a circulative propagative manner by the corn planthopper, Peregrinus maidis. MMV replicates in the nucleus of plant and insect hosts, and it remains unknown whether MMV co-opts P. maidis XPO1 (PmXPO1) to complete its life cycle. Because XPO1 plays multiple regulatory roles in cell functions and virus infection, we hypothesized that RNAi-mediated silencing of XPO1 would negatively affect MMV accumulation and insect physiology. Although PmXPO1 expression was not modulated during MMV infection, PmXPO1 knockdown negatively affected MMV accumulation in P. maidis at 12 and 15 days after microinjection. Likewise, PmXPO1 knockdown negatively affected P. maidis survival and reproduction. PmXPO1 exhibited tissue-specific expression patterns with higher expression in the ovaries compared with the guts of adult females. Survival rate was significantly lower for PmXPO1 knockdown females, compared with controls, but no effect was observed for males. PmXPO1 knockdown experiments revealed a role for PmXPO1 in ovary function and egg production. Oviposition and egg hatch on plants were dramatically reduced in females treated with dsRNA PmXPO1. These results suggest that PmXPO1 is a positive regulator of P. maidis reproduction and that it plays a proviral role in the insect vector supporting MMV infection.
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Proteína Exportina 1 , Hemípteros , Insectos Vectores , Carioferinas , Ovario , Interferencia de ARN , Receptores Citoplasmáticos y Nucleares , Animales , Femenino , Hemípteros/virología , Hemípteros/genética , Hemípteros/crecimiento & desarrollo , Carioferinas/metabolismo , Carioferinas/genética , Ovario/virología , Ovario/metabolismo , Ovario/crecimiento & desarrollo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Insectos Vectores/virología , Insectos Vectores/genética , Rhabdoviridae/fisiología , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Zea mays/virología , Zea mays/genética , Técnicas de Silenciamiento del GenRESUMEN
The endocrine system plays a pivotal role in shaping the mechanisms that ensure successful reproduction. With over a million known insect species, understanding the endocrine control of reproduction has become increasingly complex. Some of the key players include the classic insect lipid hormones juvenile hormone (JH) and ecdysteroids, and neuropeptides such as insulin-like peptides (ILPs). Individual endocrine factors not only modulate their own target tissue but also play crucial roles in crosstalk among themselves, ensuring successful vitellogenesis and oogenesis. Recent advances in omics, gene silencing, and genome editing approaches have accelerated research, offering both fundamental insights and practical applications for studying in-depth endocrine signaling pathways. This review provides an updated and integrated view of endocrine factors modulating vitellogenesis and oogenesis in insect females.
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Oogénesis , Vitelogénesis , Animales , Femenino , Insectos , Hormonas Juveniles/metabolismo , Sistema Endocrino/metabolismoRESUMEN
We describe the ultrastructure of the female reproductive organs of Deronectes moestus (Dytiscidae Hydroporinae). The long spermathecal duct has a simple epithelium lined internally by a thin cuticle and externally by a thick layer of muscle cells. The wide duct lumen contains electron-dense material, among which remnants of extracellular material are visible. This material consists of tubular structures assembled around sperm bundles previously described in the male deferent ducts. The so-called gland, disposed along the spermathecal duct, is a structure with epithelial cells lined by an irregular cuticle bearing a rich system of microvilli. Many mitochondria are visible in the apical cytoplasm of the epithelial cells, and a few spheroidal bodies are close to the basal nuclei. Since the epithelial ultrastructure of the gland suggests it is involved in fluid uptake from the lumen rather than secretory activity, the term gland, coined by other authors to describe this organ, is inappropriate. The spermatheca is a large structure with a complex epithelium showing secretory and duct-forming cells. The lumen of this organ contains sperm with the distinctive ultrastructural features of those described in the male deferent ducts, namely having a mitochondrial matrix with a small crystallized area and electron-dense dots. Because to its overall organization, the spermatheca of D. moestus can be considered a more integrated organ than those in previously studied hydroporine species.
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Escarabajos , Masculino , Animales , Femenino , Semen , Espermatozoides/ultraestructura , Células Epiteliales , Epitelio/ultraestructuraRESUMEN
The use of imidacloprid is a common pest control practice in the Neotropical region. However, the imidacloprid unintended sublethal effects on Neotropical aquatic non-target arthropods and undesirable consequences for aquatic environments remain unclear. Here, we assessed the susceptibility of Chironomus columbiensis (Diptera: Chironomidae) larvae to the neonicotinoid imidacloprid and evaluated whether sublethal exposure types would trigger sex-dependent adaptive responses (e.g., emergence, body mass, reproduction, wing morphology). We conducted a concentration-mortality curve (96 h of exposure) and established chronic and acute sublethal exposure bioassays. While chronic sublethal exposures consisted of exposing individuals during their entire larval and pupal stages, the acute sublethal exposures represented a single short duration (24 h) exposure episode during either the first or fourth larval instar. Our results revealed that chronic sublethal exposure reduced the body mass of males, while acute sublethal exposures during the first instar resulted in heavier males than those that were not exposed to imidacloprid. Chronic exposure also reduced the reproduction of males and females, while the acute sublethal exposure only affected the reproduction of individuals that were imidacloprid-exposed on their later larval instar. Chronic and acute sublethal exposures did differentially affect the wing properties of C. columbiensis males (e.g., increased size when chronically exposed and highly asymmetric wings when acutely exposed in early larval phase) and females (e.g., highly asymmetric wings when chronically and acutely exposed). Collectively, our findings demonstrated that imidacloprid can cause unintended sublethal effects on C. columbiensis, and those effects are dependent on sex, exposure type, and developmental stage.
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Chironomidae , Insecticidas , Humanos , Animales , Masculino , Femenino , Insecticidas/toxicidad , Neonicotinoides/toxicidad , Nitrocompuestos/toxicidad , LarvaRESUMEN
In anautogenous mosquitoes, synchronous development of terminal ovarian follicles after a blood meal provides an important model for studies on insect reproduction. Removal and implantation of ovaries, in vitro culture of dissected tissues and immunological assays for vitellogenin synthesis by the fat body showed that the Aedes aegypti (L.) (Diptera, Culicidae) mosquito ovary produces a factor essential for egg production. The discovery that the ovarian factor was the insect steroid hormone, ecdysone, provided a model for co-option of the larval hormones as reproductive hormones in adult insects. In later work on cultured mosquito cells, ecdysone was shown to arrest the cell cycle, resulting in an accumulation of diploid cells in G1, prior to initiation of DNA synthesis. Some mosquito species, such as Culex pipiens L. (Diptera, Culicidae), harbor the obligate intracellular bacterium, Wolbachia pipientis Hertig (Rickettsiales, Anaplasmataceae), in their reproductive tissues. When maintained in mosquito cell lines, Wolbachia abundance increases in ecdysone-arrested cells. This observation facilitated the recovery of high levels of Wolbachia from cultured cells for microinjection and genetic manipulation. In female Culex pipiens, it will be of interest to explore how hormonal cues that support initiation and progression of the vitellogenic cycle influence Wolbachia replication and transmission to subsequent generations via infected eggs.
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Diapause is a pre-programmed arrest of development allowing insects to survive in unfavorable environments. In adult insects, diapause termination is often followed by a reallocation of macronutrients and a transition to reproduction, and in some insects, this transition can be achieved using narcosis with CO2. However, whether CO2narcosis and diapause act in concert to affect reproduction remains unknown. Here, we investigated the separated and combined effects of diapause and CO2on female reproduction in queens of the common eastern bumble bee Bombus impatiens. Queens were treated with CO2 following a cold storage period (zero days, two weeks, two and four months) and were compared with untreated queens at the same timepoints for survival, colony initiation, egg-laying latency, and offspring production. We found that both CO2 and a period of at least two months in cold storage induced a transition to egg laying in gynes, and as expected, survival decreased with cold storage length. When CO2 and cold storage were combined, CO2narcosis positively affected egg laying in the earlier timepoints but its impact diminished following a longer cold storage. These data suggest that the impacts of CO2narcosis and cold storage are partially additive, and application of CO2 is effective only after a short cold storage. It further demonstrates that CO2 has complex effects on insect reproduction that are independent from diapause.
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Dióxido de Carbono , Estupor , Animales , Abejas , Dióxido de Carbono/farmacología , Femenino , Genes de Insecto , Oviposición , ReproducciónRESUMEN
Lead (Pb) stress affects hormone-mediated responses (e.g., reproduction) in insects. In this study, the effects of Pb stress (12.5-50 mg Pb/kg in larval artificial diets) on the reproduction of the common cutworm Spodoptera litura (Lepidoptera: Noctuidae) were investigated after 7 generations. The results showed that Pb stress did not reduce the longevity of adult females, but 50 mg Pb/kg significantly reduced the longevity of adult males, regardless of the generation. After 50 mg Pb/kg stress for one or 7 generations, the peak time of egg-laying was delayed, and egg production and hatchability were decreased significantly. The vitellin content in eggs was significantly inhibited by Pb stress. The S. litura vitellogenin (Vg) gene promoter was cloned and analyzed. Multiple putative transcription factors were predicted for the 2321 bp Vg promoter region, including the TATA box, GATA, basic helix-loop-helix (bHLH) transcription factor, Broad-Complex (BR-C) binding sites, etc. The fragment from -2222 to -211 bp of the Vg promoter was the activation domain for Vg, whereas the region from -211 to -55 bp repressed the activity of the Vg promoter. The construct promoter (-782/+76) in Trichoplusia ni (Hi5) cells significantly improved Vg expression, which was not affected by Pb stress (1 or 10 mg/ml). Therefore, Pb stress significantly inhibited the reproduction of S. litura but not by regulating the Vg promoter.
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Contaminantes Ambientales/toxicidad , Plomo/toxicidad , Spodoptera/fisiología , Vitelogeninas/genética , Animales , Dieta , Contaminantes Ambientales/metabolismo , Femenino , Larva/efectos de los fármacos , Plomo/metabolismo , Longevidad , Masculino , Mariposas Nocturnas , Oviposición/efectos de los fármacos , Reproducción/efectos de los fármacos , Spodoptera/efectos de los fármacos , Vitelogeninas/metabolismoRESUMEN
The ammonium transporter (AMT)/methylammonium permease (MEP)/Rhesus glycoprotein (Rh) family of ammonia (NH3/NH4+) transporters has been identified in organisms from all domains of life. In animals, fundamental roles for AMT and Rh proteins in the specific transport of ammonia across biological membranes to mitigate ammonia toxicity and aid in osmoregulation, acid-base balance, and excretion have been well documented. Here, we observed enriched Amt (AeAmt1) mRNA levels within reproductive organs of the arboviral vector mosquito, Aedes aegypti, prompting us to explore the role of AMTs in reproduction. We show that AeAmt1 is localized to sperm flagella during all stages of spermiogenesis and spermatogenesis in male testes. AeAmt1 expression in sperm flagella persists in spermatozoa that navigate the female reproductive tract following insemination and are stored within the spermathecae, as well as throughout sperm migration along the spermathecal ducts during ovulation to fertilize the descending egg. We demonstrate that RNA interference (RNAi)-mediated AeAmt1 protein knockdown leads to significant reductions (â¼40%) of spermatozoa stored in seminal vesicles of males, resulting in decreased egg viability when these males inseminate nonmated females. We suggest that AeAmt1 function in spermatozoa is to protect against ammonia toxicity based on our observations of high NH4+ levels in the densely packed spermathecae of mated females. The presence of AMT proteins, in addition to Rh proteins, across insect taxa may indicate a conserved function for AMTs in sperm viability and reproduction in general.
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Aedes/metabolismo , Compuestos de Amonio/metabolismo , Proteínas Portadoras/metabolismo , Proteínas de Insectos/metabolismo , Mosquitos Vectores/metabolismo , Amoníaco/metabolismo , Animales , Vectores de Enfermedades , Fertilidad/fisiología , Fertilización/fisiología , Masculino , ARN Mensajero/metabolismo , Reproducción/fisiología , Espermatogénesis/fisiología , Espermatozoides/metabolismo , Testículo/metabolismoRESUMEN
Oviposition is an important reproductive behavior that is triggered by mating in insects, and biogenic amines might be involved in its regulation. The effects of biogenic amines on oviposition have only been studied in a few insect species, and the findings to date have not been conclusive. In addition, there are few studies on the effects of biogenic amines on oviposition of the diamondback moth, Plutella xylostella L. Here, we tested how mating and biogenic amines regulate oviposition of P. xylostella by injecting amines and amine receptor antagonists into virgin and mated females and counting the number of eggs laid afterward. Biogenic amines of octopamine and tyramine could induce virgin adults of P. xylostella to lay eggs, while dopamine and serotonin had no such effect on oviposition. Furthermore, the octopamine antagonists mianserin, epinastine, and phentolamine inhibited oviposition by mated females. The tyramine antagonist yohimbine, dopamine antagonist SCH23390, and serotonin antagonist ketanserin did not block oviposition by mated females, and octopamine and tyramine-inducing oviposition by virgin females could be inhibited by the octopamine antagonists mianserin and epinastine instead of the tyramine antagonist yohimbine. We conclude that octopamine and its receptors are involved in mating-triggered oviposition in P. xylostella, while tyramine acts as a subsidiary. Further, the inducing effect of tyramine on oviposition is achieved via octopamine receptors instead of tyramine receptors. This experiment is helpful to further understand the role of biogenic amines in mating regulation and to provide a new strategy for controlling P. xylostella.
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BACKGROUND: Successful mating of female mosquitoes typically occurs once, with the male sperm being stored in the female spermatheca for every subsequent oviposition event. The female spermatheca is responsible for the maintenance, nourishment, and protection of the male sperm against damage during storage. Aedes aegypti is a major vector of arboviruses, including Yellow Fever, Dengue, Chikungunya, and Zika. Vector control is difficult due to this mosquito high reproductive capacity. RESULTS: Following comparative RNA-seq analyses of spermathecae obtained from virgin and inseminated females, eight transcripts were selected based on their putative roles in sperm maintenance and survival, including energy metabolism, chitin components, transcriptional regulation, hormonal signaling, enzymatic activity, antimicrobial activity, and ionic homeostasis. In situ RNA hybridization confirmed tissue-specific expression of the eight transcripts. Following RNA interference (RNAi), observed outcomes varied between targeted transcripts, affecting mosquito survival, egg morphology, fecundity, and sperm motility within the spermathecae. CONCLUSIONS: This study identified spermatheca-specific transcripts associated with sperm storage in Ae. aegypti. Using RNAi we characterized the role of eight spermathecal transcripts on various aspects of female fecundity and offspring survival. RNAi-induced knockdown of transcript AeSigP-66,427, coding for a Na+/Ca2+ protein exchanger, specifically interfered with egg production and reduced sperm motility. Our results bring new insights into the molecular basis of sperm storage and identify potential targets for Ae. aegypti control.
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Aedes/genética , Copulación , Genes de Insecto/fisiología , Inseminación , Mosquitos Vectores/genética , Motilidad Espermática , Animales , Femenino , Fertilidad/genética , Técnicas de Silenciamiento del Gen , Masculino , Interferencia de ARN , RNA-Seq , Espermatozoides/fisiología , TranscriptomaRESUMEN
Ample sperm production is essential for successful male reproduction in many species. The amount of sperm a male can produce is typically constrained by the size of his testes, which can be energetically expensive to grow and maintain. Although the economics of ejaculate allocation has been the focus of much theoretical and empirical literature, relatively little attention has been paid to individual adult variation and plasticity at the source of sperm production, the testes themselves. We experimentally address this issue using the insect Narnia femorata Stål (Hemiptera: Coreidae). We established the metabolic cost of testicular tissue and then quantified variation in individual testes mass in response to multiple mate quality and quantity treatments. We uncovered extreme variation across individuals and considerable short-term effects of mating activity on testes dry mass. Importantly, the observed variation in testes mass was associated with notable fitness consequences; females paired with males with larger testes had greater hatching success. Overall, pairing with a female resulted in a 11% reduction in dry testes mass. Despite this apparent considerable mating investment, we found no evidence of strategic allocation to higher quality females or longer-term changes in testes mass. The dynamic nature of testes mass and its metabolic cost is vital to consider in the context of re-mating rates, polyandry benefits and general mating system dynamics both in this species and more broadly.
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Hemípteros/anatomía & histología , Hemípteros/fisiología , Animales , Femenino , Masculino , Conducta Sexual Animal/fisiología , Testículo/anatomía & histologíaRESUMEN
Vitellogenesis is pre-requisite to insect egg production and embryonic development after oviposition. During insect vitellogenesis, the yolk protein precursor vitellogenin (Vg) is mainly synthesized in the fat body, transported by the hemolymph through the intercellular spaces (known as patency) in the follicular epithelium to reach the membrane of maturing oocytes, and sequestered into the maturing oocytes via receptor-mediated endocytosis. Insect vitellogenesis is governed by two critical hormones, the sesquiterpenoid juvenile hormone (JH) and the ecdysteriod 20-hydroxyecdysone (20E). JH acts as the principal gonadotropic hormone to stimulate vitellogenesis in basal hemimetabolous and most holometabolous insects. 20E is critical for vitellogenesis in some hymenopterans, lepidopterans and dipterans. Furthermore, microRNA (miRNA) and nutritional (amino acid/Target of Rapamycin and insulin) pathways interplay with JH and 20E signaling cascades to control insect vitellogenesis. Revealing the regulatory mechanisms underlying insect vitellogenesis is critical for understanding insect reproduction and helpful for developing new strategies of insect pest control. Here, we outline the recent research progress in the molecular action of gonadotropic JH and 20E along with the role of miRNA and nutritional sensor in regulating insect vitellogenesis. We highlight the advancements in the regulatory mechanisms of insect vitellogenesis by the coordination of hormone, miRNA and nutritional signaling pathways.
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Lepidopteran species present an interesting case of sperm polymorphism and testicular fusion. The study of these features are of great importance in understanding the reproductive biology of these insects, especially in the case of those considered pests. Dione juno and Agraulis vanillae stand out as the most important pests of passion fruit (Passiflora sp.) crops in Brazil. Therefore, the objective of the present study was to characterize the testes and germ cells of Dione juno and Agraulis vanillae at different life stages, using light microscopy and scanning and transmission electron microscopy, to understand the maturation mechanisms of the male gametes in these species. The study showed that the larvae of both species have a pair of brown kidney-shaped testes, covered by epithelial cells which divide the organ into four follicles. The testes are full of spermatogonia which begin to differentiate in the third larval instar. In the fifth larval instar, spermatozoa can be observed. When they enter the prepupal stage the testes begin a fusion process that is completed in the adult insects, where they present as spherical organs divided into eight follicles, containing all the cells of the germ line. Spermatogenesis occurs centripetally, and in both species, sperm dimorphism is observed, where two different types of spermatozoa are formed, eupyrene (nucleated) and apyrene (anucleate), which differ in morphology and function. Apart from contributing to scientific basic research on the reproductive biology of these insects, the present study provides important data that can aid in research on the physiology, systematics, and control of these species.
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Mariposas Diurnas , Espermatogénesis/fisiología , Espermatogonias/citología , Testículo/anatomía & histología , Testículo/ultraestructura , Animales , Brasil , Mariposas Diurnas/anatomía & histología , Mariposas Diurnas/fisiología , Mariposas Diurnas/ultraestructura , Masculino , Microscopía Electrónica de Rastreo , Microscopía Electrónica de Transmisión , Passiflora/parasitologíaRESUMEN
The springtail Collembola are characterized by having rolled spermatozoa, with a long cylindrical extracellular structure adhering to the acrosome. This structure is produced by the secretory activity of the testes epithelial cells at almost the end of spermiogenesis. At the beginning of its formation, it is a thin extension with a helical wall and a dense axial region. Later the cylindrical structure shows an inner organization which is different in the several species examined: species of Entomobryidae contain material with a paracrystalline structure, whilst some of Symphypleona contain ovoid structures. The outer envelope of the extracellular structure consists of two overlapped layers orthogonally arranged, clearly identified by cryo-preparations. Immunoblot analysis and lectin stainings have indicated that the cylindrical structure has a glycoproteic composition. As the structure is no longer visible after the sperm transfer into the female spermatheca, it is suggested that it could contain enzymes able to activate the sperm unwinding process and possibly allowing the reacquisition of sperm motility.
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Acrosoma/química , Acrosoma/ultraestructura , Artrópodos/citología , Glicoproteínas/análisis , Complejos Multiproteicos/análisis , Espermatozoides/química , Espermatozoides/citología , Animales , Immunoblotting , Masculino , Coloración y EtiquetadoRESUMEN
Vitellogenin (Vg), a yolk protein precursor, is the primary egg nutrient source involved in insect reproduction and embryo development. The Cotton Boll weevil (CBW) Anthonomus grandis Boheman, the most important cotton pest in Americas, accumulates large amounts of Vg during reproduction. However, the precise role of this protein during embryo development in this insect remains unknown. Herein, we investigated the effects of vitellogenin (AgraVg) knockdown on the egg-laying and egg viability in A. grandis females, and also characterized morphologically the unviable eggs. AgraVg transcripts were found during all developmental stages of A. grandis, with highest abundance in females. Silencing of AgraVg culminated in a significant reduction in transcript amount, around 90%. Despite this transcriptional reduction, egg-laying was not affected in dsRNA-treated females but almost 100% of the eggs lost their viability. Eggs from dsRNA-treated females showed aberrant embryos phenotype suggesting interference at different stages of embryonic development. Unlike for other insects, the AgraVg knockdown did not affect the egg-laying ability of A. grandis, but hampered A. grandis reproduction by perturbing embryo development. We concluded that the Vg protein is essential for A. grandis reproduction and a good candidate to bio-engineer the resistance against this devastating cotton pest.
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Long-chain acyl-CoA esters are important intermediates in lipid metabolism and are synthesized from fatty acids by long-chain acyl-CoA synthetases (ACSL). The hematophagous insect Rhodnius prolixus, a vector of Chagas' disease, produces glycerolipids in the midgut after a blood meal, which are stored as triacylglycerol in the fat body and eggs. We identified twenty acyl-CoA synthetase genes in R. prolixus, two encoding ACSL isoforms (RhoprAcsl1 and RhoprAcsl2). RhoprAcsl1 transcripts increased in posterior midgut on the second day after feeding, and RhoprAcsl2 was highly transcribed on the tenth day. Both enzymes were expressed in Escherichia coli. Recombinant RhoprACSL1 and RhoprACSL2 had broad pH optima (7.5-9.5 and 6.5-9.5, respectively), were inhibited by triacsin C, and were rosiglitazone-insensitive. Both showed similar apparent Km for palmitic and oleic acid (2-6 µM), but different Km for arachidonic acid (0.5 and 6 µM for RhoprACSL1-Flag and RhoprACSL2-Flag, respectively). The knockdown of RhoprAcsl1 did not result in noticeable phenotypes. However, RhoprACSL2 deficient insects exhibited a 2.5-fold increase in triacylglycerol content in the fat body, and 90% decrease in fatty acid ß-oxidation. RhoprAcsl2 knockdown also resulted in 20% increase in lifespan, delayed digestion, 30% reduced oviposition, and 50% reduction in egg hatching. Laid eggs and hatched nymphs showed remarkable alterations in morphology. In summary, R. prolixus ACSL isoforms have distinct roles on lipid metabolism. Although RhoprACSL1 functions remain unclear, we propose that RhoprACSL2 is the main contributor for the formation of the intracellular acyl-CoA pool channeled for ß-oxidation in the fat body, and is also required for normal reproduction.