Cypermethrin induction of vitellogenesis and ovarian development in unfed adult female Ornithodoros moubata (Acari: Argasidae)

Summary

Vitellogenesis in ticks is known to be induced by engorgement and mating. In this paper, the synthetic pyrethroid cypermethrin (CyM) is shown to induce production of yolk protein precursor, vitellogenin (Vg), and ovarian development in unengorged mated adult female Ornithodoros moubata. The levels of Vg found in the hemolymph and ovarian development induced by CyM were dose-dependent. i.e., CyM doses of more than 0.2 and 1.0 μg/tick were needed for significant increase of Vg titer in the hemolymph and yolk deposition in oocytes, respectively. Immunological and electrophoretical analyses of Vg and Vitellin (Vn) induced by CyM were identical with those induced by engorgement. Vg titer induced by CyM in unengorged females followed approximately the same time course as that in the normal engorged females. However, Vg titer induced by CyM continued to increase after day 8 and reached a maximum (95 μg/μ1) on day 10 after treatment, while Vg titer induced by engorgement decreased again after reaching a maximum (60 μg/μ1) on day 6, correlated with yolk Vn deposition in oocytes. Ovarian development induced by even high doses (10 or 20 μg/tick) of CyM was slow compared to normal development stimulated by engorgement. Oviposition was not observed in females treated with CyM.     

Suppression of allatotropin simulates reproductive diapause in the mosquito Culex pipiens

The cessation of juvenile hormone (JH) production is a key endocrine event that halts ovarian development and hence initiates diapause in females of the mosquito, Culex pipiens. The shutdown in endocrine activity of the corpora allata (CA), the source of JH, was manifested in the smaller size of CA in females reared under short daylengths (diapause) compared to those reared under long daylengths (nondiapause), as well as in low expression of the mRNA encoding allatotropin, the neuropeptide that promotes JH biosynthesis in the CA. Genes encoding both allatotropin and allatostatin were identified in C. pipiens, but only expression levels of allatotropin differed in the two types of females. Knockdown of allatotropin mRNA using RNA interference in females programmed for nondiapause resulted in a cessation of ovarian development akin to diapause. This arrest in development could be reversed with an application of JH. Our results thus suggest that suppression of allatotropin is a critical link in regulating the shutdown of the CA during diapause.     

Cyanoprotein: Developmental stage,sex and diapause-dependent expression,and synthesis regulation by juvenile hormone in the bean bug,Riptortus clavatus

Cyanoprotein (CP) synthesis was studied in nymphal and nondiapause adult, diapause adult, and juvenile hormone (JH) treated adult bean bugs, Riptortus clavatus. Hemolymph collected from bugs injected with [³⁵S]-methionine was analyzed by native polyacrylamide gel electrophoresis (PAGE) and fluorography, and CP synthesis was also determined quantitatively by rocket immunoelectrophoresis of hemolymph and counting. In the nymphal stages synthesis of CP-1, 2, 3, and 4 (with CP-4 synthesis predominant) reached a maximum at mid-instar and was not detected at each ecdysis, so that the synthetic activity of CPs changed cyclically in each instar. During the nymphal stages CP synthesis showed the same pattern and level in both females and males, and both diapause and nondiapause oriented bugs. In the adult stage, however, CP synthesis differed in the two sexes and nondiapause or diapause conditions. In nondiapause males CP synthesis was not detected in the adult, but in nondiapause females CP (only CP-1) was synthesized through the reproductive stages. CP-1 accumulated in the egg yolk together with vitellogenin. In diapause adults (both females and males) CP-1 to 4 were synthesized at a very low rate for over 2 months and accumulated in the hemolymph. JH or JH analog (JHA) methoprene and also long day condition, which terminate diapause, switched the main CP synthesis CP-4 to CP-1 in females. CP synthesis in diapause males stopped after JH(A) treatment. The activities of CP synthesis, thus, changed in developmental stages, sexes, and diapause. This is an excellent system for study of specific gene expression and switching controlled by insect hormones and sex. © 1992 Wiley-Liss, Inc.     

Imaginal induction of diapause in several 'adult-female diapausing' spider mites

Abstract. The possibility of imaginal induction of diapause in adult female spider mites is suggested in Stigmaeopsis miscanthi. Here, several spider mites are studied to investigate the generality of this phenomenon. The existence of imaginal induction of reproductive diapause is established in Stigmaeopsis longus, two forms of S. miscanthi and Tetranychus kanzawai adult females. The females that develop under diapause-averting (long days and 18 °C) conditions are induced to diapause under short-daylength conditions at 18 °C after moulting to the adult stage. However, the adult females of Tetranychus urticae never diapause under these conditions.     

Both the anterior and posterior eyes function as photoreceptors for photoperiodic termination of diapause in the two-spotted spider mite

Photoreceptors involved in photoperiodism in insects and mites can be either the retinal photoreceptors in the visual system or nonvisual extraretinal photoreceptors. Mites with no eyes have a clear photoperiodic response, suggesting the involvement of extraretinal photoreceptors in mite photoperiodism. In mites equipped with eyes, however, it is not known whether the retinal or extraretinal photoreceptors are involved in photoperiodism. The two-spotted spider mite Tetranychus urticae possesses two pairs of eyes. Adult females of this species terminate diapause in response to long days. To investigate whether the eyes function as photoperiodic photoreceptors in T. urticae, their eyes were ablated using a laser ablation system. Mites with their eyes intact terminated diapause under long days after low temperature exposure, whereas they remained in diapause under short days. Under constant darkness, they did not terminate diapause. When all eyes were removed, the mites remained in diapause even when they were maintained under long days. In contrast, the mites showed clear photoperiodic response when only the anterior or posterior eyes were removed. These results indicate that both the anterior and posterior eyes function as photoreceptors in photoperiodic termination of diapause in T. urticae.     

Molecular analysis of nutritional and hormonal regulation of female reproduction in the red flour beetle, Tribolium castaneum

Female reproduction includes maturation of oocytes and the synthesis of yolk proteins (vitellogenin, Vg) in the fat body and their deposition into the oocytes. Our recent studies showed that juvenile hormone (JH) regulates Vg synthesis and 20-hydroxyecdysone (20E) regulates oocyte maturation in the red flour beetle (Tribolium castaneum). Here, we report on the role of nutritional signaling on vitellogenesis and oogenesis. Comparison of gene expression between fed and starved beetles by microarray analysis showed the up-regulation of genes involved in energy homeostasis and down-regulation of genes involved in egg production in the starved beetles. The RNA interference (RNAi) aided knock-down in the expression of genes involved in insulin and TOR signaling pathways showed that both these signaling pathways play key roles in Vg synthesis and oocyte maturation. Starvation of female beetles resulted in a block in Vg synthesis but not in the progression of primary oocyte development to the resting stage. Feeding after starvation induced Vg synthesis and the progression of primary oocytes from the resting stage to the mature stage. However, in the beetles where JH or 20E synthesis or action was blocked by RNAi, both Vg synthesis and oocyte maturation were affected suggesting that both these hormones (JH and 20E) and nutritional signaling and their cross-talk regulate vitellogenesis and oogenesis.     

Genome wide gene-expression analysis of facultative reproductive diapause in the two-spotted spider mite Tetranychus urticae

Background

Diapause or developmental arrest, is one of the major adaptations that allows mites and insects to survive unfavorable conditions. Diapause evokes a number of physiological, morphological and molecular modifications. In general, diapause is characterized by a suppression of the metabolism, change in behavior, increased stress tolerance and often by the synthesis of cryoprotectants. At the molecular level, diapause is less studied but characterized by a complex and regulated change in gene-expression. The spider mite Tetranychus urticae is a serious polyphagous pest that exhibits a reproductive facultative diapause, which allows it to survive winter conditions. Diapausing mites turn deeply orange in color, stop feeding and do not lay eggs.

Results

We investigated essential physiological processes in diapausing mites by studying genome-wide expression changes, using a custom built microarray. Analysis of this dataset showed that a remarkable number, 11% of the total number of predicted T. urticae genes, were differentially expressed. Gene Ontology analysis revealed that many metabolic pathways were affected in diapausing females. Genes related to digestion and detoxification, cryoprotection, carotenoid synthesis and the organization of the cytoskeleton were profoundly influenced by the state of diapause. Furthermore, we identified and analyzed an unique class of putative antifreeze proteins that were highly upregulated in diapausing females. We also further confirmed the involvement of horizontally transferred carotenoid synthesis genes in diapause and different color morphs of T. urticae.

Conclusions

This study offers the first in-depth analysis of genome-wide gene-expression patterns related to diapause in a member of the Chelicerata, and further adds to our understanding of the overall strategies of diapause in arthropods.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-14-815) contains supplementary material, which is available to authorized users.     

Localization of vitellogenin mRNA expression and vitellogenin uptake during ovarian maturation in the giant freshwater prawn Macrobrachium rosenbergii

In situ hybridization and immunohistochemical techniques were used to investigate the dynamics of vitellogenin (Vg) mRNA expression and Vg uptake during ovarian maturation in the hepatopancreas and ovary at differing stages of ovarian maturation in both intact and eyestalk ablated female Macrobrachium rosenbergii. In the hepatopancreas of intact animals, Vg mRNA expression was detected faintly two days after ecdysis, and signals showed a gradual increase as the molt cycle advanced to the premolt stages, but decreased at the late premolt stage. Vg mRNA was detected in the R-cells of the hepatopancreas, indicating that these cells are responsible for synthesizing Vg. No Vg mRNA expression was observed in the ovary. Immunohistochemistry results for the hepatopancreas showed a pattern of staining intensity similar to that of in situ hybridization. Increases in the accumulation of yolk protein in the oocytes occurred concomitantly with increasing Vg mRNA expression. In eyestalk ablated animals, Vg mRNA expression and Vg uptake showed similar but accelerated patterns to those of intact animals. This study has confirmed on the cellular level previous results that Vg synthesis is intrinsically correlated to ovarian maturation and the molt cycle in M. rosenbergii.     

Effects of acclimation and diapause on the thermal tolerance of the two-spotted spider mite Tetranychus urticae

The two-spotted spider mite, Tetranychus urticae, is a worldwide pest species that overwinters as diapausing females. Cold hardening is presumed to start during diapause development to ensure the successful overwintering of this species. To address this hypothesis, we compared cold tolerance between non-diapausing and diapausing females. We measured supercooling point (SCP) and survival to acute cold stress by exposing the mites at a range of sub-zero temperatures (from −4 to −28 °C for 2 h). The mean SCPs of non-diapausing and diapausing females were −19.6±0.5 and −24.7±0.3 °C respectively, and freezing killed the mites. Diapausing females were significantly more cold tolerant than non-diapausing ones, with LT₅₀ of −19.7 and −13.3 °C, respectively. Further, we also examined the effects of cold acclimation (10 d at 0 or 5 °C) in non-diapausing and diapausing females. Our findings indicated that diapause decreased SCP significantly, while cold acclimation had no effect on the SCP except for non-diapausing females that were acclimated at 5 °C. Acclimation at 5 °C enhanced survival to acute cold stress in diapausing and non-diapausing females, with LT₅₀ of −22.0 and −17.1 °C, respectively. Altogether, our results indicate that T. urticae is a chill tolerant species, and that diapause and cold acclimation elevate cold hardiness in this species.     

The effect of temperature and light conditions on diapause induction in a Korean population of Neoseiulus womersleyi Schicha (Acari: Phytoseiidae)

To determine the condition for diapause induction of a Korean population of Neoseiulus womersleyi, combinations of constant temperatures (14, 16, 18, 20 and 25 °C) and photoperiods (0, 8, 10, 12, 14, 16 and 24 h photophase) were used from egg to adult emergence. Diapause induction was determined by reproductive cessation of adult females. Lower temperature and shorter photophase resulted in higher diapause induction. Critical photophases for diapause induction were 10 and 12 h at 18 and 16 °C, respectively. Diapause-induced N. womersleyi adult females consumed significantly fewer eggs of the two-spotted spider mite, Tetranychus urticae, than non-induced females. Field monitoring of N. womersleyi showed that there was no egg-bearing female after 282 Julian date, while a model estimated complete diapause induction at 288 Julian date. Diapause of this predatory mite occurred approximately 2 weeks before diapause of its main prey, T. urticae, in Korean apple orchards. Further study perspectives are discussed relative to the spider mite biological control system in fruit orchards.     

Antigenic and electrophoretic variants of vitellogenin in Oncopeltus blood and their control by juvenile hormone

Antigenic analysis of adult female-specific blood and yolk proteins in Oncopeltus demonstrated an incomplete vitellogenin (A), which appears in the blood prior to yolk deposition and is later modified or joined by an antigenically complete molecule (AB). Vitellogenin AB is antigenically indistinguishable from the major yolk protein of mature eggs, though the electrophoretic mobilities of the two differ in 6% acrylamide gels. Vitellogenin A alone appears in the blood of adult females in which the corpora allata are known to be inactive, i.e., during starvation or photoperiodically induced diapause. Stimulation of these females with a juvenile hormone analog restores yolk deposition, and also induces the appearance of AB in the blood. While juvenile hormone is needed for the termination of diapause and the maturation of vitellogenin in this species, diapause begins with the vitellogenin-producing mechanism already partially assembled.     

Regulation of vitellogenin synthesis by juvenile hormone analogue in Coccinella septempunctata

Vitellogenesis in the lady beetle, Coccinella septempunctata, is controlled by juvenile hormone (JH). When immature females were reared on an artificial diet, they were characterized by hypertrophy of the fat bodies and slow development of the ovaries. Vitellogenin (Vg) synthesis in the fat body remained at a very low level throughout adult development. RNA synthesis also stayed at a relatively low level. Treatment with hydroprene induced vitellogenesis in these non-fecund females. Stimulation in Vg synthesis in hormone-treated females was demonstrated both in vivo and in fat body culture. Synthesis of fat body RNA also increased after hormone treatment. Fat body RNA from hormone-treated females directed the synthesis of Vg polypeptides both in Xenopus oocytes and in a reticulocyte lysate. Thus the induction of Vg synthesis by JH involves an increase in the level of translatable Vg mRNA.     

Inheritance of the photoperiodic response controlling imaginal summer diapause in the cabbage beetle, Colaphellus bowringi

Adults of the cabbage beetle Colaphellus bowringi display a summer diapause in response to the exposure of their larvae to long photoperiods. In the present study, the inheritance of the photoperiodic response controlling summer diapause in C. bowringi by crossing a high diapause strain (D strain) with a laboratory selected nondiapause strain (N strain) was investigated under different photoperiods at 22, 25 and 28 °C. The beetles in both reciprocal crosses and backcrosses showed a clear short-day response for the induction of diapause at all temperatures, similar to that of the D strain, suggesting that photoperiodic response of this beetle is heritable. The diapause incidences in the progeny from all the crosses under LD 15:9 or LD 12:12 at 25 °C suggest that genetic and genetic-environmental interactions are involved in diapause induction. The incidence of diapauses in F₁ progeny was significantly lower than that in the D × D strain but significantly higher than that in the N × N strain, indicating that the diapause capability is inherited in an incomplete dominant manner. The incidence of diapause was greater among the offspring of hybrid females when those females had a D strain mother or grandmother rather than a N strain mother or grandmother, indicating that maternal effects on diapause induction are stronger than paternal effects. The laboratory selected nondiapause strain also showed a short-day photoperiodic response at a low temperature of 22 °C, indicating that the photoperiodic photoreceptor and photoperiodic clock still function in the nondiapause strain.     

Dynamics of vitellogenin mRNA expression during vitellogenesis in the banana shrimp Penaeus (Fenneropenaeus) merguiensis using real-time PCR

An open reading frame (ORF) of vitellogenin (Vg) cDNA was amplified from the ovaries of the banana shrimp, Penaeus merguiensis. An examination of Vg-deduced amino acid sequence revealed the presence of cleavage sites at a consensus motif for subtilisin-like endoproteases prior to the N-terminal sequences of purified vitellin (Vt) subunits. A comparison of the primary structures of Vg molecules in decapod crustacean species revealed the existence of a common characteristic structure, and phylogenetic analysis reflected the current taxonomic classifications of crustaceans. A PCR product of 1.1 kb encoding the 3'-end of Vg cDNA was cloned from the hepatopancreas. Although its sequence was almost identical to that of the same region of the ovarian Vg, with only 18 nucleotide differences, analysis suggests that they have been subjected to natural selection, indicating that there may be two different, tissue-specific Vg genes in P. merguiensis. This is consistent with the different expression patterns of Vg mRNA, as determined by real-time PCR. Vg mRNA levels were maintained at low levels during the previtellogenic stage and they increased as vitellogenesis progressed to reach a peak at the early vitellogenic stage in the ovary or at the vitellogenic stage in the hepatopancreas, and thereafter, levels decreased. Expression of Vg mRNA was much higher in the ovary compared to the hepatopancreas at all stages of ovarian development, implying that the ovary is mainly responsible for Vt synthesis. These indicate that penaeids constitute a unique model for vitellogenesis, showing intraovarian gene expression and synthesis of yolk protein.     

Drosophila americana Diapausing Females Show Features Typical of Young Flies

Diapause is a period of arrested development which is controlled physiologically, preprogrammed environmentally and characterized by metabolic depression that can occur during any stage of insect development. Nevertheless, in the genus Drosophila, diapause is almost always associated with the cessation of ovarian development and reproductive activity in adult females. In this work, we show that, in D. americana (a temperate species of the virilis group), diapause is a genetically determined delay in ovarian development that is triggered by temperature and/or photoperiod. Moreover, we show that in this species diapause incidence increases with latitude, ranging from 13% in the southernmost to 91% in the northernmost range of the distribution. When exposed to diapause inducing conditions, both diapausing and non-diapausing females show a 10% increase in lifespan, that is further increased by 18.6% in diapausing females, although senescence is far from being negligible. ActinD1 expression levels suggest that diapausing females are biologically much younger than their chronological age, and that the fly as a whole, rather than the ovarian development alone, which is phenotypically more evident, is delayed by diapause. Therefore, diapause candidate genes that show expression levels that are compatible with flies younger than their chronological age may not necessarily play a role in reproductive diapause and in adaptation to seasonally varying environmental conditions.     

Maintenance of diapause variability in the two-spotted spider mite,tetranychus urticae, in a heterogeneous and stochastic environment

A simple nongenetic mathematical model analyzed the processes responsible for the variations in the diapause percentage among populations of the two-spotted spider mite,Tetranychus urticae Koch. This model incorporates the following assumptions. 1) Mites have diapause (DD), non-diapause (NN), and “plastic” (DN) populations (plasticity exists in the phenotypic expression of diapause in response to habitat conditions at the time of diapause induction). 2) A heterogeneous mite habitat consists of microhabitat L, in which all the non-diapausing mites die during the winter due to the lack of winter host plants, and microhabitat O with winter hosts capable of supporting some of the non-diapausing mites overwinter. 3) Temporal fluctuation of winter conditions which affect the survival and reproduction of non-diapausing mites. Using these assumptions, we compared the fitness functions of the three populations and analyzed the conditions under which each population is favoured over the other two, thereby elucidating the processes involved in the maintenance of variability in diapause. Our analysis revealed: 1) frequent mild winters are of primary importance for the non-diapause trait to be maintained, 2) the existence of winter hosts is also important for the nondiapause trait to be favoured, and this importance depends greatly on the degree of the adaptive diapause expression in the DN mites in response to habitat conditions, i.e., the better the phenotypeenvironment matching in DN, the higher the probability that DN will be favoured, 3) The combined effect of the temporal and spatial variation enhances the maintenance of variablity in the diapause trait of the mites.     

Expression of heat shock protein 70a mRNA in Bombyx mori diapause eggs

In an effort to understand whether heat shock protein 70 (Hsp70) participates in the environmental 5 °C signal reception/transduction toward breaking embryonic diapause of the silkworm Bombyx mori, we isolated a cDNA for Hsp70a and examined the expression of Hsp70a mRNA in B. mori diapause and nondiapause eggs by quantitative real-time PCR. Hsp70a mRNA gradually increased in diapause eggs continuously kept at 25 °C after oviposition to maintain diapause. When diapause eggs were exposed to the diapause-terminating condition of 5 °C beginning at 2 days post-oviposition, Hsp70a mRNA increased beginning at 5 days post-cold treatment. Even in nondiapause eggs, Hsp70a mRNA increased slightly with exposure to 5 °C. These results suggest that Hsp70a is involved in reception/transduction of the diapause-terminating (5 °C) signal via gene activation. The expression patterns of Hsp70a mRNA are discussed in relation to those of the cold-response gene Samui.