Application Of Juvenile Hormone Research Articles

The BTB transcription factor, Abrupt, acts cooperatively with Chronologically inappropriate morphogenesis (Chinmo) to repress metamorphosis and promotes leg regeneration

Many insects undergo the process of metamorphosis when larval precursor cells begin to differentiate to create the adult body. The larval precursor cells retain stem cell-like properties and contribute to the regenerative ability of larval appendages. Here we demonstrate that two Broad-complex/Tramtrack/Bric-à-brac Zinc-finger (BTB) domain transcription factors, Chronologically inappropriate morphogenesis (Chinmo) and Abrupt (Ab), act cooperatively to repress metamorphosis in the flour beetle, Tribolium castaneum. Knockdown of chinmo led to precocious development of pupal legs and antennae. We show that although topical application of juvenile hormone (JH) prevents the decrease in chinmo expression in the final instar, chinmo and JH act in distinct pathways. Another gene encoding the BTB domain transcription factor, Ab, was also necessary for the suppression of broad (br) expression in T. castaneum in a chinmo RNAi background, and simultaneous knockdown of ab and chinmo led to the precocious onset of metamorphosis. Furthermore, knockdown of ab led to the loss of regenerative potential of larval legs independently of br. In contrast, chinmo knockdown larvae exhibited pupal leg regeneration when a larval leg was ablated. Taken together, our results show that both ab and chinmo are necessary for the maintenance of the larval tissue identity and, apart from its role in repressing br, ab acts as a crucial regulator of larval leg regeneration. Our findings indicate that BTB domain proteins interact in a complex manner to regulate larval and pupal tissue homeostasis.

Juvenile Hormone-Induced Abnormality of <i>Periplaneta americana</i> (L.)

Application of juvenile hormone III (JH III) and a JH analogue, methoprene to last instars of Periplaneta americana suppressed adult emergence and caused morphological abnormalities such as twisted wings and asymmetry of appendages resulting in formation of either nymphoids or giant supernumerary nymphs and adultoids. The effects of JH III and methoprene were concentration dependent in inducing aberrations in morphogenetic moult as well as in significantly lowering the body mass (BM) and lipid reserves (LR) in P. americana.

Transcriptomics on Social Interactions in Termites: Effects of Soldier Presence

The organization of social insect colonies requires sophisticated mechanisms to regulate caste composition according to colony demands. In termites, the soldier caste is responsible for the inhibition of soldier differentiation, but the mechanism underlying the regulation of soldier differentiation is still unclear. In this study, we performed transcriptome analyses to identify genes expressed in workers that fluctuated in the presence of soldiers in the subterranean termite Reticulitermes speratus. First, soldier differentiation was artificially induced via juvenile hormone (JH) application, and the inhibitory effects of soldier differentiation on soldier presence were evaluated. Second, transcriptomes were prepared from workers with or without soldiers under JH treatment, and expression analyses were performed to identify differentially expressed genes (DEGs) for each treatment. The expression levels of several DEGs were verified by quantitative real-time PCR. The results indicated that only a small number of DEGs were upregulated by the presence of soldiers. A homology search of DEGs and gene ontology (GO) analysis of the DEGs showed that some genes were responsible for the regulation of hormone levels, social interaction, and response to xenobiotic substances, suggesting that they could be involved in developmental arrest and pheromonal regulation in workers. Moreover, GO analysis indicated that the expression of many genes, including those involved in hormone metabolic processes, fluctuated with JH application. Suppression of soldier differentiation in the presence of soldiers could be accomplished by the expression of a large number of genes required for soldier differentiation.

Juvenile hormone as a causal factor for maternal regulation of diapause in a wasp

Most temperate multivoltine insects enter diapause, a hormonally controlled developmental suspension, in response to seasonal photoperiodic and/or thermal cues. Some insect species exhibit maternal regulation of diapause in which developmental trajectories of the offspring are determined by mothers in response to environmental cues that the mother received. Although maternally regulated diapause is common among insects, the maternal endocrinological mechanisms are largely veiled. To approach this issue, we used the jewel wasp Nasonia vitripennis, which produces non-diapause-destined offspring under long days and diapause-destined offspring under short days or low temperatures. Comparative transcriptomics of these wasps revealed possible involvement of the juvenile hormone (JH) biosynthetic cascade in maternal diapause regulation. The expression of juvenile hormone acid O-methyltransferase (jhamt) was typically downregulated in short-day wasps, and this was reflected by a reduction in haemolymph JH concentrations. RNAi targeted at jhamt reduced haemolymph JH concentration and induced wasps to produce diapause-destined offspring even under long days. In addition, topical application of JH suppressed the production of diapause-destined offspring under short days or low temperatures. These results indicate that diapause in N. vitripennis is determined by maternal jhamt expression and haemolymph JH concentration in response to day length. We therefore report a novel role for JH in insect seasonality.

Juvenile hormone regulation of microRNAs is mediated by E75 in the Dengue vector mosquito Aedes aegypti

MicroRNAs (miRNAs) are small noncoding RNAs that play critical roles in controlling posttranscriptional gene regulation and have a profound effect on mosquito reproduction and metabolism. Juvenile hormone (JH) is critical for achieving reproductive competence in the main vector of human arboviral diseases, Aedes aegypti We report a JH-mediated mechanism governing miRNA expression. Using a transcription factor screen with multiple primary miRNA (pri-miRNA) promoters, we identified that the Ecdysone-induced protein E75 (E75) isoform (E75-RD) induced miRNA gene promoter activity. E75 binding sites were determined in miRNA promoters by means of cell transfection assay. E75-RD was found to be up-regulated by JH, as shown by the JH application and RNA interference (RNAi) of the JH receptor Methoprene-tolerant (Met). Small RNA sequencing from RNAi of Met and E75 displayed an overlapping miRNA cohort, suggesting E75 to be an intermediate component within the JH hierarchical network controlling miRNAs. Further experiments confirmed that E75-RD positively regulates several miRNAs including miR-2940. Reducing miR-2940 resulted in the arrest of follicle development and number of eggs laid. Performing miRNA target predictions and RT-qPCR from antagomir Ant-2940-3p-treated fat body tissues identified the mRNA target Clumsy (AAEL002518) The molecular interaction between this gene target and miR-2940 was confirmed using an in vitro dual luciferase assay in Drosophila S2 cells and in Ae. aegypti Aag2 cell lines. Finally, we performed a phenotypic rescue experiment to demonstrate that miR-2940/Clumsy is responsible for the disruption in egg development. Collectively, these results established the role of JH-mediated E75-RD in regulation of miRNA gene expression during the mosquito reproductive cycle.

Monoterpenoid signals and their transcriptional responses to feeding and juvenile hormone regulation in bark beetle Ips hauseri.

Hauser's engraver beetle, Ips hauseri, is a serious pest in spruce forest ecosystems in Central Asia. Its monoterpenoid signal production, transcriptome responses and potential regulatory mechanisms remain poorly understood. The quality and quantity of volatile metabolites in hindgut extracts of I. hauseri were found to differ between males and females and among three groups: beetles that were newly emerged, those with a topical application of juvenile hormone III (JHIII) and those that had been feeding for 24 h. Feeding males definitively dominated monoterpenoid signal production in I. hauseri, which uses (4S)-(-)-ipsenol and (S)-(-)-cis-verbenol to implement reproductive segregation from Ipstypographus and Ipsshangrila. Feeding stimulation induced higher expression of most genes related to the biosynthesis of (4S)-(-)-ipsenol than JHIII induction, and showed a male-specific mode in I. hauseri. JHIII stimulated males to produce large amounts of (-)-verbenone and also upregulated the expression of several CYP6 genes, to a greater extent in males than in females. The expression of genes involved in the metabolism of JHIII in females and males was also found to be upregulated. Our results indicate that a species-specific aggregation pheromone system for I. hauseri, consisting of (4S)-(-)-ipsenol and S-(-)-cis-verbenol, can be used to monitor population dynamics or mass trap killing. Our results also enable a better understanding of the bottom-up role of feeding behaviors in mediating population reproduction/aggregation and interspecific interactions.

Ovicidal activity of juvenile hormone mimics in the bean bug, Riptortus pedestris.

Insect juvenile hormone (JH) mimics (JHMs) are known to have ovicidal effects if applied to adult females or eggs. Here, we examined the effects of exogenous JHMs on embryonic development of the bean bug, Riptortus pedestris. The expression profiles of JH early response genes and JH biosynthetic enzymes indicated that JH titer was low for the first 3 days of the egg stage and increased thereafter. Application of JH III skipped bisepoxide (JHSB3) or JHM on Day 0 eggs when JH titer was low caused reduced hatchability, and the embryos mainly arrested in mid- or late embryonic stage. Application of JHMs on Day 5 eggs also resulted in an arrest, but this was less effective compared with Day 0 treatment. Interestingly, ovicidal activity of synthetic JHMs was much lower than that of JHSB3. This study will contribute to developing novel insecticides that are selective among insect species.

Effect of Larval Topical Application of Juvenile Hormone on Cuticular Chemical Composition of Mischocyttarus consimilis (Vespidae: Polistinae) Females

Juvenile Hormone (JH) is considered the main determinant of caste in social insects, though little is known about how this hormone acts in social wasps, especially the independent-founding species. The known relationship between JH titer and caste in the colony and we suggest a relationship among the effects of JH and the cuticular chemical profile. Therefore, this study aimed to test the hypothesis that topical application of JH to larvae of different instars alters the cuticular chemical composition of newly emerged females of Mischocyttarus consimilis (Zikán), influencing the dynamics of colony. Two techniques were used to evaluate the variation in cuticular chemical composition: Fourier transform infrared photoacoustic spectroscopy and gas chromatography coupled to mass spectrometry (GC-MS). Indeed, the application of JH did significantly alter the cuticular chemical composition of adult females that received treatment at the larval stage in comparison to control. The effects of JH were instar-dependent in that the results of topical application were significant when performed at third larval instar. Overall, these results add evidence that caste determination may, at least in part may be pre-imaginal in species of independent-founding social wasps.

Methyl farnesoate regulatory mechanisms underlying photoperiod‐dependent sex determination in the freshwater crustacean Daphnia magna

Freshwater zooplankton Daphnia magna has been widely used in ecotoxicology studies. During the last 20 years, it has been demonstrated that the topical application of juvenile hormone (JH) or JH analogs to mother daphnids induce male offspring production. Based on this finding, an in vivo screening validation method for chemicals with JH agonistic effect has developed. Although this screening system successfully identified a number of JH-like chemicals, molecular mechanisms underlying the male sex-determining process remain largely unknown. To address this issue, we established a reliable male- or female-producing system using Daphnia pulex WTN6 strain by changing the rearing photoperiod. Taking advantage of this rearing system, we successfully found several factors involving male sex determination such as ionotropic glutamate receptors, protein kinase C and pantothenate. Here, we used two D. magna strains that can also control the production of female or male offspring by photoperiod differences as model species for ecotoxicology studies. We demonstrated that either treatment of antagonist of ionotropic glutamate receptors or inhibitor of protein kinase C strongly suppressed male offspring production even under male-producing conditions. Moreover, we revealed that male sex-determining processes are likely diverged between D. magna and D. pulex based on the current experiment. This study provides a fine experimental method for in vivo screening not only JH agonists but also JH antagonists. Moreover, using daphnids with photoperiod-dependent sex determination manner will hugely contribute to understanding the mode-of-action of JH in daphnids.

The POU factor Ventral veins lacking regulates ecdysone and juvenile hormone biosynthesis during development and reproduction of the milkweed bug, Oncopeltus fasciatus

Recent studies have demonstrated endocrine roles for the POU domain transcription factor Ventral veins lacking (Vvl) during larval development of holometabolous insects - insects that undergo complete metamorphosis. In this study, the role of Vvl was examined in the milkweed bug, Oncopeltus fasciatus, a hemimetabolous insect. In the embryos, vvl was found to be expressed in the presumptive prothoracic glands. When vvl expression was knocked down using RNA interference (RNAi), embryos arrested their development after dorsal closure. Vvl double-stranded RNA (dsRNA)-injected nymphs failed to molt and had reduced expression of the ecdysone response gene, hormone receptor 3 (HR3), the ecdysone biosynthesis genes, disembodied and spook, and the juvenile hormone (JH) response gene, Krüppel homolog 1 (Kr-h1). Injection of 20-hydroxyecdysone rescued the molting phenotype and HR3 expression in vvl knockdown nymphs. In adults, vvl RNAi inhibited egg laying and suppressed the expression of Kr-h1 and vitellogenin in the fat body. Application of JH III or methoprene restored oviposition in vvl knockdown adults, indicating that Vvl regulates JH biosynthesis during reproduction. Thus, Vvl functions as a critical regulator of hormone biosynthesis throughout all developmental stages of O. fasciatus. Our study demonstrates that Vvl is a critical transcription factor involved in JH and ecdysteroid biosynthesis in both hemimetabolous and holometabolous insects.

Insulin signaling mediates previtellogenic development and enhances juvenile hormone-mediated vitellogenesis in a lepidopteran insect, Maruca vitrata

BackgroundInsulin/insulin-like growth peptide signaling (IIS) down-regulates hemolymph sugar level and facilitates larval growth in the soybean pod borer, Maruca vitrata. The objective of this study is to determine whether IIS of M. vitrata can mediate ovarian development of adult females.ResultsA pair of ovaries consists of 8 ovarioles, each of which is separated into distal germarium and proximal vitellarium in M. vitrata. In the germarium, oocyte development occurred with active mitotic activity which was visible by incorporating bromodeoxyribose uridine. Previtellogenic development and subsequent vitellogenesis began soon after adult emergence. They continued with increase of female age. Oocyte development was facilitated by up-regulation of vitellogenin (Vg) and Vg receptor (VgR) gene expression. Larval diets significantly influenced on ovarian development of M. vitrata because oocyte development varied with pupal size derived from larvae treated with different nutritional diets. Its ovarian development was dependent on endocrine signal(s) from the head because decapitation soon after adult emergence prevented oogenesis and subsequent vitellogenesis along with marked reduction of Vg and VgR expression. Topical application of juvenile hormone (JH) significantly recovered its ovarian development whereas farnesoic acid (a precursor of JH biosynthesis) or 20-hydroxyecdysone treatment did not. JH stimulated vitellogenesis and choriogenesis, but not previtellogenic development. In contrast, insulin injection to decapitated females stimulated oocyte differentiation and vitellogenesis along with increase of Vg and VgR expression. To further analyze the effect of insulin on ovarian development, expression of four IIS components (InR, FOXO, Akt, and TOR) genes was manipulated by RNA interference. Hemocoelic injection of gene-specific double stranded RNAs significantly reduced their target gene mRNA levels and interfered with ovarian development. An addition of insulin to JH treatment against decapitated females enhanced the gonadotropic effect of JH by stimulating oogenesis.ConclusionsIIS plays crucial role in mediating previtellogenic development of M. vitrata in response to nutrient signal. It also enhances the gonadotropic effect of JH II on vitellogenesis.

MiR-34 modulates wing polyphenism in planthopper.

Polyphenism is a successful strategy adopted by organisms to adapt to environmental changes. Brown planthoppers (BPH, Nilaparvata lugens) develop two wing phenotypes, including long-winged (LW) and short-winged (SW) morphs. Though insulin receptor (InR) and juvenile hormone (JH) have been known to regulate wing polyphenism in BPH, the interaction between these regulators remains largely elusive. Here, we discovered that a conserved microRNA, miR-34, modulates a positive autoregulatory feedback loop of JH and insulin/IGF signaling (IIS) pathway to control wing polyphenism in BPH. Nlu-miR-34 is abundant in SW BPHs and suppresses NlInR1 by targeting at two binding sites in the 3’UTR of NlInR1. Overexpressing miR-34 in LW BPHs by injecting agomir-34 induces the development towards SW BPHs, whereas knocking down miR-34 in SW BPHs by injecting antagomir-34 induces more LW BPHs when another NlInR1 suppressor, NlInR2, is also suppressed simultaneously. A cis-response element of Broad Complex (Br-C) is found in the promoter region of Nlu-miR-34, suggesting that 20-hydroxyecdysone (20E) might be involved in wing polyphenism regulation. Topic application of 20E downregulates miR-34 expression but does not change wing morphs. On the other hand, JH application upregulates miR-34 expression and induces more SW BPHs. Moreover, knocking down genes in IIS pathway changes JH titers and miR-34 abundance. In all, we showed that miRNA mediates the cross talk between JH, 20E and IIS pathway by forming a positive feedback loop, uncovering a comprehensive regulation mechanism which integrates almost all known regulators controlling wing polyphenism in insects.

Effects of larval Age at Grafting and Juvenile Hormone on Morphometry and Reproductive Quality Parameters of in Vitro Reared Honey Bees (Hymenoptera: Apidae).

The honey bee queen plays a central role in the Apis mellifera L. (Hymenoptera: Apidae) colony, and her high reproductive capacity is fundamental for building up the workforce of a colony. Caste development in honey bee females involves elaborate physiological pathways unleashed at the beginning of the first larval instars, with juvenile hormone (JH) playing a crucial role. Here we took advantage of established in vitro rearing techniques to conduct a 2 × 2 experimental design and test initial rearing age (young vs old) and JH treatment (JH III vs solvent control) to enlighten the role of nutrient quality and JH in shaping honey bee female fertility, morphological features related to queenliness, and key physiological parameters (hemolymph vitellogenin/Vg, sugar levels, and Vg transcript levels). Our results show that while the age at initial larval rearing had major impacts on external morphology development, where younger larvae exhibited a higher probability to develop into queen-like adults morphotypes, the JH application during the larval stage improved physiological pathways related to ovary development and metabolism during the ontogenic development. We detected that the supplementation of queen larvae with JH promoted important benefits regarding queen fertility as the increase of ovariole number and vg levels at hemolymph, both crucial factors at eggs production. The data presented here provide guidance in efforts to improve honey bee queen quality, especially in light of frequent episodes of queen failures in the beekeeping industry.

Vitamin D1 versus ecdysteroids: Growth effects on cell regeneration and malignant growth in insects are similar to those in humans

Polyhydroxylated derivatives of 6-keto,7-dehydrocholesterol (ecdysteroids) are common constituents of various plants. In 1965, they were accidentally discovered in the search for the insect moulting hormone. These biologically important natural compounds are neither insect hormones nor inducers of insect ecdysis. Due to their strong anabolic, vitamin D-like effects in insects, domestic animals and humans, I propose the use of the arbitrary term vitamin D1. The present paper describes the effects of vitamin D1 on the growth and regeneration of excised epidermal cells of the tobacco hornworm, Manduca sexta (Sphingidae). The periods of programmed cell death and cell proliferation (histolysis and histogenesis, respectively) exactly coincide in insects with endogenous peaks of increased concentration of vitamin D1. Epidermal cells communicate with each other, creating a mutually integrated tissue, connected by mechanical, chemical, electrical, ionic or other so far incompletely known factors. After natural cell death, or after the artificial removal of some epidermal cells, the neighbouring cells that lose communication integrity, begin to divide mitotically to replace the disconnected part. Cell divisions are arrested as soon as the integrity of the living tissue is established. During insect ontogeny, the application of juvenile hormone causes regenerating epidermal cells to repeat the previous morphogenetic programme (i.e., development of patches of larval tissue on the body of a pupa, or metathetely). Conversely, the application of vitamin D1 (20-hydroxyecdysone) caused the regenerating cells to prematurely execute a future morphogenetic programme (i.e., development of patches of pupal tissue on the body of a larva, or prothetely). Among the key features of insect regeneration, is the arrest of cell divisions when tissues resume living cell-to-cell integrity. This prevents the formation of aberrant groups of cells, or tumours. It is well established that the main physiological systems of insects (e.g., circulatory, respiratory, neuro-endocrine) are structurally and functionally similar to corresponding systems in humans. Thus the basic principles of cell regeneration and the role of vitamin D1 in insects may also be valid for humans. The common vitamins D2 (ergocalciferol) or D3 (cholecalciferol), are exclusively lipid soluble secosterols, which require activation by UV irradiation and hydroxylation in the liver. By contrast, the neglected vitamin D1 is a natural derivative of polyhydroxylated 7-dehydrocholesterol of predominantly plant origin, which is both partly a water and partly a lipid soluble vitamin. It neither requires UV irradiation, nor hydroxylation due to 6 or 7 already built-in hydroxylic groups. Like other vitamins, it enters insect or human bodies in plant food or is produced by intestinal symbionts. Vitamin D1 causes strong anabolic, vitamin D-like effects in domestic animals and in humans. I am convinced that avitaminosis associated with a deficiency of vitamin D1 in human blood may be responsible for certain hitherto incurable human diseases, especially those related to impaired nerve functions and somatic growth, aberrant cell regeneration or formation of malignant tumours.

Experimental improvement of honey bee (Apis mellifera) queen quality through nutritional and hormonal supplementation

Queen reproductive potential (=quality) impacts the health and productivity of honey bee colonies. To determine the factors that affect reproductive quality during development, we tested queens produced under larval treatments by supplementing the diet with juvenile hormone (JH), additional sugars, or both, compared to untreated control. Furthermore, we varied the age of the larvae that were grafted (1 and 3 days old). We analyzed newly emerged virgin queens for their morphological characters as proxies for their reproductive potential. We found that the application of a sugar-enriched diet in combination with JH application onto 1st instar queen larvae produced higher-quality queens, while for 3rd instar larvae only the JH treatment resulted in increasing queen quality. For mated queens, those treated with JH plus supplemented sugars showed a significantly higher sperm count and sperm viability. Our findings demonstrate that honey bee queen reproductive potential can be increased through diet supplementation.

Cloning and characterization of Methoprene-tolerant (Met) and Krüppel homolog 1 (Kr-h1) genes in the wheat blossom midge, Sitodiplosis mosellana.

Juvenile hormone (JH), a growth regulator, inhibits ecdysteroid-induced metamorphosis and controls insect development and diapause. Methoprene-tolerant (Met) and Krüppel homolog 1 (Kr-h1) are two proteins involved in JH action. To gain some insight into their function in development of Sitodiplosis mosellana, an insect pest undergoing obligatory larval diapause at the mature 3rd instar stage, we cloned full-length complementary DNAs of Met and Kr-h1 from this species. SmMet encoded a putative protein, which contained three domains typical of the bHLH-PAS family and eight conserved amino acid residues important for JH binding. SmKr-h1 encoded a protein showing high sequence homology to its counterparts in other species, and contained all eight highly conserved Zn-finger motifs for DNA-binding. Expression patterns of SmMet and SmKr-h1 were developmentally regulated and JH III responsive as well. Their mRNA abundance increased as larvae entered early 3rd instar, pre-diapause and maintenance stages, and peaked during post-diapause quiescence, a pattern correlated with JH titers in this species. Different from reduced expression of SmMet, SmKr-h1 mRNA increased at mid-to-late period of post-diapause development. Topical application of JH III on diapausing larvae also induced the two genes in a dose-dependent manner. Expression of SmMet and SmKr-h1 clearly declined in the pre-pupal phase, and was significantly higher in female adults than male adults. These results suggest that JH-responsive SmMet and SmKr-h1 might play key roles in diapause induction and maintenance as well as in post-diapause quiescence and adult reproduction, whereas metamorphosis from larvae to pupae might be correlated with their reduced expression.

Juvenile hormone mediates the positive effects of nitrogen fertilization on weight and reproduction in pea aphid.

The positive effects of nitrogen fertilization on the performance of phytophagous insects have been reported extensively; the physiological and molecular basis involved, however, is largely unclear. Here, we test experimentally whether enhancement of juvenile hormone (JH) is responsible for the increased weight and fecundity of pea aphid (Acyrthosiphon pisum) under nitrogen fertilization. Aphids fed on Medicago truncatula with nitrogen fertilization have a greater amino acid content, higher weight at the fourth instar and adult stage, and produce more offspring than those without nitrogen fertilization. Furthermore, nitrogen fertilization upregulates the transcripts of JH biosynthesis-related genes and increases JH titre at the fourth instar and adult stage, suggesting that JH is involved in the positive responses of aphids to nitrogen fertilization. Application of 100 ng JH increases adult weight and fecundity in aphids fed on M. truncatula without nitrogen fertilization. Conversely, impairing JH signalling by pharmacologically inhibiting the target of rapamycin pathway or by knocking down JH biosynthetic gene decreases adult weight and fecundity in aphids fed on M. truncatula with nitrogen fertilization, whereas application of JH rescued the phenotype. The increased JH titre at the fourth instar and adult stage is required for the increases of weight and fecundity of A. pisum under nitrogen fertilization. © 2018 Society of Chemical Industry.

Bactericera cockerelli vitellogenin-6 like, a vitellogenin without a direct reproductive function?

Vitellogenin-like proteins are members of the large lipid transfer proteins, a family of proteins involved in reproduction, lipid circulation and immune defences. In this study, we identified a new Bactericera cockerelli vitellogenin-like (Vg-like) transcript, and named it BcVg6-like based on its similarity to Acyrthosiphon pisum Vg6. In silico analyses predicted different conserved domains in BcVg6-like compared with the conventional Ba.cockerelli vitellogenin, BcVg1-like, previously described by our research group. Phylogenetic analyses determined that BcVg6-like clustered with Vg-like-B proteins and not the conventional vitellogenins involved in vitellogenesis. Also, the expression analyses showed differences in BcVg6-like transcript expression between 7-day-old males and 3- and 7-day-old females. BcVg6-like was not upregulated after exogenous application of juvenile hormone III, but its relative expression increased significantly in alimentary canals of adult females exposed to tomato plants infected by the bacterial plant pathogen 'Candidatus Liberibacter solanacearum'. Our results suggest that in Ba.cockerelli, both vitellogenin genes may have different functions: BcVg1-like is a conventional vitellogenin that conserved its ancestral function as an egg yolk precursor whereas BcVg6-like might have acquired a function in lipid and/or other molecule transport, and could potentially play a role in immune defence.

Distinct Roles of Met and Interacting Proteins on the Expressions of takeout Family Genes in Brown Planthopper.

The takeout family genes encode relatively small proteins that are related to olfaction and are regulated by juvenile hormone (JH). The takeout genes modulate various physiological processes, such as behavioral plasticity in the migratory locust Locusta migraloria and feeding and courtship behaviors in Drosophila. Therefore, to understand the regulatory mechanism of these physiological processes, it is important to study the expressions of the takeout genes that are regulated by JH signaling. We used quantitative real-time PCR (qRTPCR) to study the role of JH signaling in the regulation of the takeout family genes in the brown planthopper Nilaparvata lugens (N. lugens) through the application of Juvenile hormone III (JHIII) and the down-regulation of key genes in the JH signaling pathway. The topical application of JHIII induced the expressions of most of the takeout family genes, and their expressions decreased 2 and 3 days after the JHIII application. Down-regulating the brown planthopper JH receptor NlMethoprene-tolerant (NlMet) and its interacting partners, NlTaiman (NlTai) and Nlß-Ftz-F1 (Nlß-Ftz), through RNAi, exhibited distinct effects on the expressions of the takeout family genes. The down-regulation of NlMet and NlKrüppel-homolog 1 (NlKr-h1) increased the expressions of the takeout family genes, while the down-regulation of the Met interacting partners NlTai and Nlß-Ftz decreased the expressions of most of the takeout family genes. This work advanced our understanding of the molecular function and the regulatory mechanism of JH signaling.

Identification and expression analyses of vitellogenin in Bactericera cockerelli (Šulc)

The potato psyllid, Bactericera cockerelli (Šulc) (Hemiptera: Triozidae), is a phloem-feeding insect with preference for Solanaceae. This insect species is vector of the pathogenic bacteria ‘Candidatus Liberibacter solanacearum’ the causative agent of zebra chip, an important disease of commercial potatoes in several countries worldwide. The recent classification of psyllids among the most dangerous vectors has promoted their study, but still many biological processes such as reproduction and vitellogenesis need to be investigated. As a first step towards the elucidation of vitellogenesis in B. cockerelli, one candidate vitellogenin transcript (6622 bases long) was identified and the expression of the transcript and the protein were analyzed in virgin and mated females between 1 and 7days old. In virgin females, Vg expression increased up to 5days old; while mating significantly up-regulated Vg expression in 5- and 7-day-old females. To determine the role of juvenile hormone in B. cockerelli Vg expression, topical applications of juvenile hormone III were performed on virgin females, resulting in an up-regulation of Vg expression and an increase in the number of mature oocytes present in female reproductive organs. Overall, this study represents the first step to understand vitellogenesis of B. cockerelli and it highlights the role of JH III in the hormonal regulation of Vg expression and oocyte development.