A fatty acid anabolic pathway in specialized-cells sustains a remote signal that controls egg activation in Drosophila.

Egg activation, representing the critical oocyte-to-embryo transition, provokes meiosis completion, modification of the vitelline membrane to prevent polyspermy, and translation of maternally provided mRNAs. This transition is triggered by a calcium signal induced by spermatozoon fertilization in most animal species, but not in insects. In Drosophila melanogaster, mature oocytes remain arrested at metaphase-I of meiosis and the calcium-dependent activation occurs while the oocyte moves through the genital tract. Here, we discovered that the oenocytes of fruitfly females are required for egg activation. Oenocytes, cells specialized in lipid-metabolism, are located beneath the abdominal cuticle. In adult flies, they synthesize the fatty acids (FAs) that are the precursors of cuticular hydrocarbons (CHCs), including pheromones. The oenocyte-targeted knockdown of a set of FA-anabolic enzymes, involved in very-long-chain fatty acid (VLCFA) synthesis, leads to a defect in egg activation. Given that some but not all of the identified enzymes are required for CHC/pheromone biogenesis, this putative VLCFA-dependent remote control may rely on an as-yet unidentified CHC or may function in parallel to CHC biogenesis. Additionally, we discovered that the most posterior ventral oenocyte cluster is in close proximity to the uterus. Since oocytes dissected from females deficient in this FA-anabolic pathway can be activated in vitro, this regulatory loop likely operates upstream of the calcium trigger. To our knowledge, our findings provide the first evidence that a physiological extra-genital signal remotely controls egg activation. Moreover, our study highlights a potential metabolic link between pheromone-mediated partner recognition and egg activation.

[Azadirachtin, a natural pesticide with multiple effects]. / L'azadirachtine, un pesticide naturel aux effets multiples.

There are many studies devoted to the negative impact of conventional pesticides that effectively control pests, but cause widespread environmental pollution. As a result, interest is growing in pesticides of a natural origin with a lower environmental impact. Among them, azadirachtin, sold under various formulations (neem oil, Neem-Azal, Bioneem, etc.), is still the most widely recommended molecule in agricultural ecosystems. Azadirachtin has also been used in traditional medicine for centuries, and studies published over the past few years have tended to support its therapeutic use. Yet the argument that azadirachtin is harmless to the environment has been offset by its notable collateral and controversial effects on non-target organisms. The present paper summarizes the work already done in this field.

TITLE: L'azadirachtine, un pesticide naturel aux effets multiples. ABSTRACT: Une littérature abondante traite de l'impact négatif des pesticides conventionnels, très efficaces dans la gestion des ravageurs mais responsables d'une large pollution environnementale. Les pesticides d'origine naturelle qui auraient un moindre impact environnemental suscitent ainsi un intérêt majeur. Parmi ceux-ci, l'azadirachtine, commercialisée sous diverses formulations (huile de neem, Neem-Azal, Bioneem, etc.) reste la molécule la plus recommandée dans les agro-écosystèmes. L'argument d'une innocuité environnementale de l'azadirachtine est cependant nuancé par des effets collatéraux qui, bien que controversés, sont notables sur des organismes non ciblés.

Control of pheromone production by ovaries in Drosophila.

Drosophila pheromones are long chain hydrocarbons (CHCs) produced by specialized epidermal cells, the oenocytes. Here we were explored the role of ovaries in CHC regulation. We studied tudor, a grandchildless-like mutation, resulting in progeny without ovaries and three alleles of ovoD, resulting in ovarian defects depending on the strength of the allele. We show here that these mutant flies with no or abnormal ovaries have a decrease in C29 length CHC ratio, balanced by an increase in C23 and C25 length ratio; this effect is dependent on the strength of the mutation. An increase in the amount of CHCs also occurred but was not related to the strength of ovoD alleles. As ovaries are the main site of ecdysone production in females, we knocked down the receptor to ecdysone EcR in the oenocytes and obtained increased amounts of CHCs and inhibition of long chain CHC synthesis, showing that the lack of an ecdysone signal arriving into the oenocytes is responsible for these defects. We then investigated the role of follicular cells and oocyte on CHC regulation: we RNAi-knocked down the LPR receptors in the oocyte to hinder vitellogenesis without significantly modifying CHC profile. We then expressed apoptosis genes within the follicle cells or within the ovocytes and obtained less long chain and more short chain CHC levels in the former case and an enhanced CHC production in the latter case. Together, these results support the notion of an interaction between oocyte and follicular cells, which send an ecdysone signal to the oenocytes to regulate CHC synthesis.

Azadirachtin effects on mating success, gametic abnormalities and progeny survival in Drosophila melanogaster (Diptera).

BACKGROUND: Azadirachtin is a prominent natural pesticide and represents an alternative to conventional insecticides. It has been successfully used against insect pests. However, its effects on reproduction require further analysis. Here we investigated lethal and sublethal effects of azadirachtin, on treated adults in a model insect, Drosophila melanogaster (Meigen). Dose-mortality relationships as well as several parameters of reproduction (mating, spermatogenesis, oogenesis and fertility) were examined. RESULTS: Neem-Azal, a commercial formulation of azadirachtin, applied topically on newly emerged adults, increased mortality with a positive dose-dependent relationship. The LD50 (0.63 µg) was determined 24 h after treatment using a non-linear regression. Adults surviving this dose had a mating success that was divided by 3 and a progeny production reduced by half when males were treated, and even more when females were treated. When combining probability of survival, of mating and reduced progeny, it appeared that LD50 induced a 98% reduction in reproductive rates. Reduced progeny was partially explained by the effect of adult treatment on gametes number and abnormalities. The number of cysts and the apical nuclei positions within the cysts decreased by 29.7% and 20%, respectively, in males. In females, the number of oocytes per ovary and the volume of basal oocytes also decreased by 16.1% and 32.4%, respectively. CONCLUSION: Azadirachtin causes significant toxic effects in both sexes and decreases the fecundity and fertility of D. melanogaster. Females are more sensitive to azadirachtin. © 2017 Society of Chemical Industry.

Male accessory gland proteins affect differentially female sexual receptivity and remating in closely related Drosophila species.

In sexual species, mating success depends on the male's capacity to find sexual partners and on female receptivity to mating. Mating is under evolutionary constraints to prevent interspecific mating and to maximize the reproductive success of both sexes. In Drosophila melanogaster, female receptivity to mating is mainly controlled by Sex peptide (SP, i.e. Acp70A) produced by the male accessory glands with other proteins (Acps). The transfer of SP during copulation dramatically reduces female receptivity to mating and prevents remating with other males. To date, female postmating responses are well-known in D. melanogaster but have been barely investigated in closely-related species or strains exhibiting different mating systems (monoandrous versus polyandrous). Here, we describe the diversity of mating systems in two strains of D. melanogaster and the three species of the yakuba complex. Remating delay and sexual receptivity were measured in cross-experiments following SP orthologs or Acp injections within females. Interestingly, we discovered strong differences between the two strains of D. melanogaster as well as among the three species of the yakuba complex. These results suggest that reproductive behavior is under the control of complex sexual interactions between the sexes and evolves rapidly, even among closely-related species.

Hydrocarbon Patterns and Mating Behaviour in Populations of Drosophila yakuba.

Drosophila yakuba is widespread in Africa. Here we compare the cuticular hydrocarbon (CHC) profiles and mating behavior of mainland (Kounden, Cameroon) and island (Mayotte, Sao-Tome, Bioko) populations. The strains each had different CHC profiles: Bioko and Kounden were the most similar, while Mayotte and Sao-Tome contained significant amounts of 7-heptacosene. The CHC profile of the Sao-Tome population differed the most, with half the 7-tricosene of the other populations and more 7-heptacosene and 7-nonacosene. We also studied the characteristics of the mating behavior of the four strains: copulation duration was similar but latency times were higher in Mayotte and Sao-Tome populations. We found partial reproductive isolation between populations, especially in male-choice experiments with Sao-Tome females.

Flexible origin of hydrocarbon/pheromone precursors in Drosophila melanogaster.

In terrestrial insects, cuticular hydrocarbons (CHCs) provide protection from desiccation. Specific CHCs can also act as pheromones, which are important for successful mating. Oenocytes are abdominal cells thought to act as specialized units for CHC biogenesis that consists of long-chain fatty acid (LCFA) synthesis, optional desaturation(s), elongation to very long-chain fatty acids (VLCFAs), and removal of the carboxyl group. By investigating CHC biogenesis in Drosophila melanogaster, we showed that VLCFA synthesis takes place only within the oenocytes. Conversely, several pathways, which may compensate for one another, can feed the oenocyte pool of LCFAs, suggesting that this step is a critical node for regulating CHC synthesis. Importantly, flies deficient in LCFA synthesis sacrificed their triacylglycerol stores while maintaining some CHC production. Moreover, pheromone production was lower in adult flies that emerged from larvae that were fed excess dietary lipids, and their mating success was lower. Further, we showed that pheromone production in the oenocytes depends on lipid metabolism in the fat tissue and that fatty acid transport protein, a bipartite acyl-CoA synthase (ACS)/FA transporter, likely acts through its ACS domain in the oenocyte pathway of CHC biogenesis. Our study highlights the importance of environmental and physiological inputs in regulating LCFA synthesis to eventually control sexual communication in a polyphagous animal.

Acp70A regulates Drosophila pheromones through juvenile hormone induction.

Mated Drosophila melanogaster females show a decrease in mating receptivity, enhanced ovogenesis, egg-laying and activation of juvenile hormone (JH) production. Components in the male seminal fluid, especially the sex peptide ACP70A stimulate these responses in females. Here we demonstrate that ACP70A is involved in the down-regulation of female sex pheromones and hydrocarbon (CHC) production. Drosophila G10 females which express Acp70A under the control of the vitellogenin gene yp1, produced fewer pheromones and CHCs. There was a dose-dependent relationship between the number of yp1-Acp70A alleles and the reduction of these compounds. Similarly, a decrease in CHCs and diene pheromones was observed in da > Acp70A flies that ubiquitously overexpress Acp70A. Quantitative-PCR experiments showed that the expression of Acp70A in G10 females was the same as in control males and 5 times lower than in da > Acp70A females. Three to four days after injection with 4.8 pmol ACP70A, females from two different strains, exhibited a significant decrease in CHC and pheromone levels. Similar phenotypes were observed in ACP70A injected flies whose ACP70A receptor expression was knocked-down by RNAi and in flies which overexpress ACP70A N-terminal domain. These results suggest that the action of ACP70A on CHCs could be a consequence of JH activation. Female flies exposed to a JH analog had reduced amounts of pheromones, whereas genetic ablation of the corpora allata or knock-down of the JH receptor Met, resulted in higher amounts of both CHCs and pheromonal dienes. Mating had negligible effects on CHC levels, however pheromone amounts were slightly reduced 3 and 4 days post copulation. The physiological significance of ACP70A on female pheromone synthesis is discussed.

[What the study of seminal fluid proteins in Drosophila tells us about the evolution of reproduction]. / Ce que l'étude des protéines du fluide séminal de drosophile nous apprend sur l'évolution de la reproduction.

Decrease in male fertility observed in the past decades have involved sperm quantity and quality disorders. However, decrease in quality or quantity of seminal fluid may also trigger drastic reduction of female and also male fertility. The present paper documents on the composition of seminal fluid, the consequences on sperm cells and on the physiological and behavioral effects towards females. The work evidences the crucial role of seminal fluid in the postcopulatory interactions between the sexes and illustrates the selective effects in the male-female coevolution.

Male pheromone polymorphism and reproductive isolation in populations of Drosophila simulans.

The dominant cuticular hydrocarbons (HC) in Drosophila simulans are 7-tricosene (7-T) and 7-pentacosene (7-P). The 7-T is the major HC in East Africa and in other continents. In West Africa, D. simulans is very rare and displays 7-P as the major compound. We studied three D. simulans strains from Egypt (Eg), Sao-Tome (ST), and Cameroon (Cam), with 7-T, intermediary or 7-P phenotypes. HC profiles of ST and Cam female differed slightly from corresponding male profiles; females had more 7-T and less 7-P. Varying temperature affected all HCs (even those with 27 and 29 carbons)-not just 7-T and 7-P; there was no clear relationship between HC phenotype and resistance to desiccation. We report reproductive isolation between Eg and ST and Eg and Cam (but not between ST and Cam), which is due to Eg and Cam female preferences for their own males. In conclusion, our findings do support divergence of D. simulans populations from West Africa for both pheromonal profile and mating preference.

Endarterectomy Versus Angioplasty in Patients with Symptomatic Severe Carotid Stenosis (EVA-3S) trial: results up to 4 years from a randomised, multicentre trial.

BACKGROUND: Carotid stenting is a potential alternative to carotid endarterectomy but whether this technique is as safe as surgery and whether the long-term protection against stroke is similar to that of surgery are unclear. We previously reported that in patients in the Endarterectomy Versus Angioplasty in Patients with Symptomatic Severe Carotid Stenosis (EVA-3S) trial, the rate of any stroke or death within 30 days after the procedure was higher with stenting than with endarterectomy. We now report the results up to 4 years. METHODS: In this follow-up study of a multicentre, randomised, open, assessor-blinded, non-inferiority trial, we compared outcome after stenting with outcome after endarterectomy in 527 patients who had carotid stenosis of at least 60% that had recently become symptomatic. The primary endpoint of the EVA-3S trial was the rate of any periprocedural stroke or death (ie, within 30 days after the procedure). The prespecified main secondary endpoint was a composite of any periprocedural stroke or death and any non-procedural ipsilateral stroke during up to 4 years of follow-up. Other trial outcomes were any stroke or periprocedural death, any stroke or death, and the above endpoints restricted to disabling or fatal strokes. This trial is registered with ClinicalTrials.gov, number NCT00190398. FINDINGS: 262 patients were randomly assigned to endarterectomy and 265 to stenting. The cumulative probability of periprocedural stroke or death and non-procedural ipsilateral stroke after 4 years of follow-up was higher with stenting than with endarterectomy (11.1%vs 6.2%, hazard ratio [HR] 1.97, 95% CI 1.06-3.67; p=0.03). The HR for periprocedural disabling stroke or death and non-procedural fatal or disabling ipsilateral stroke was 2.00 (0.75-5.33; p=0.17). A hazard function analysis showed the 4-year differences in the cumulative probabilities of outcomes between stenting and endarterectomy were largely accounted for by the higher periprocedural (within 30 days of the procedure) risk of stenting compared with endarterectomy. After the periprocedural period, the risk of ipsilateral stroke was low and similar in both treatment groups. For any stroke or periprocedural death, the HR was 1.77 (1.03-3.02; p=0.04). For any stroke or death, the HR was 1.39 (0.96-2.00; p=0.08). INTERPRETATION: The results of this study suggest that carotid stenting is as effective as carotid endarterectomy for middle-term prevention of ipsilateral stroke, but the safety of carotid stenting needs to be improved before it can be used as an alternative to carotid endarterectomy in patients with symptomatic carotid stenosis.