Anopheles gambiae mosquitoes are the principal vectors of malaria. A major determinant of the capacity of these mosquitoes as disease vectors is their high reproductive rate. Reproduction depends on a single insemination, which profoundly changes the behavior and physiology of females. To identify factors and mechanisms relevant to the fertility of A. gambiae, we performed a comprehensive analysis of the molecular and cellular machinery associated with copulation in females. Initial whole-body microarray experiments comparing virgins with females at 2 h, 6 h, and 24 h after mating detected large transcriptional changes. Analysis of tissue localization identified a subset of genes whose expression was strikingly regulated by mating in the lower reproductive tract and, surprisingly, the gut. In the atrium of virgin females, where the male seminal fluid is received, our studies revealed a ''mating machinery'' consisting of molecular and structural components that are turned off or collapse after copulation, suggesting that this tissue loses its competence for further insemination. In the sperm storage organ, we detected a number of mating-responsive genes likely to have a role in the maintenance and function of stored sperm. These results identify genes and mechanisms regulating the reproductive biology of A. gambiae females, highlighting considerable differences with Drosophila melanogaster. Our data inform vector control strategies and reveal promising targets for the manipulation of fertility in field populations of these important disease vectors.mosquito ͉ post-mating response ͉ reproduction ͉ microarray ͉ reproductive tract

Anopheles gambiae sensu stricto is the major vector of malaria, a disease with devastating consequences for human health. Given the constant spread of the disease, alternative approaches to the use of insecticides are urgently needed to control vector populations. Females of this species undergo large behavioral changes after mating, which include a life-long refractoriness to further insemination and the induction of egg laying in blood-fed individuals. Genetic control strategies aimed at impacting Anopheles fertility through the release of sterile males are being advocated to reduce the size of mosquito field populations. Such strategies depend on the ability of the released sterile males to mate successfully with wild females and to switch off the female receptivity to further copulation. Here we evaluate the role of sperm in regulating female behavioral responses after mating in An. gambiae. We developed spermless males by RNAi silencing of a germ cell differentiation gene. These males mated successfully and preserved standard accessory gland functions. Females mated to spermless males exhibited normal postcopulatory responses, which included laying large numbers of eggs upon blood feeding and becoming refractory to subsequent insemination. Moreover, spermless males induced transcriptional changes in female reproductive genes comparable to those elicited by fertile males. Our data demonstrate that, in contrast to Drosophila, targeting sperm in An. gambiae preserves normal male and female reproductive behavior for the traits and time frame analyzed and validate the use of approaches based on incapacitation or elimination of sperm for genetic control of vector populations to block malaria transmission.oviposition | reproduction | sexual refractoriness | postmating behavior | insect

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Anopheles gambiae and An. coluzzii, the two most important malaria vectors in sub-Saharan Africa, are recently radiated sibling species that are reproductively isolated even in areas of sympatry. In females from these species, sexual transfer of male accessory gland products, including the steroid hormone 20-hydroxyecdysone (20E), induces vast behavioral, physiological, and transcriptional changes that profoundly shape their post-mating ecology, and that may have contributed to the insurgence of post-mating, prezygotic reproductive barriers. As these barriers can be detected by studying transcriptional changes induced by mating, we set out to analyze the post-mating response of An. gambiae and An. coluzzii females captured in natural mating swarms in Burkina Faso. While the molecular pathways shaping short- and long-term mating-induced changes are largely conserved in females from the two species, we unravel significant inter-specific differences that suggest divergent regulation of key reproductive processes such as egg development, processing of seminal secretion, and mating behavior, that may have played a role in reproductive isolation. Interestingly, a number of these changes occur in genes previously shown to be regulated by the sexual transfer of 20E and may be due to divergent utilization of this steroid hormone in the two species.

Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) is the major Afro-tropical vector of malaria. Novel strategies proposed for the elimination and eradication of this mosquito vector are based on the use of genetic approaches, such as the sterile insect technique (SIT). These approaches rely on the ability of released males to mate with wild females, and depend on the application of effective protocols to assess the swarming and mating behaviours of laboratory-reared insects prior to their release. The present study evaluated whether large semi-field enclosures can be utilized to study the ability of males from a laboratory colony to respond to natural environmental stimuli and initiate normal mating behaviour. Laboratory-reared males exhibited spatiotemporally consistent swarming behaviour within the study enclosures. Swarm initiation, peak and termination time closely tracked sunset. Comparable insemination rates were observed in females captured in copula in the semi-field cages relative to females in small laboratory cages. Oviposition rates after blood feeding were also similar to those observed in laboratory settings. The data suggest that outdoor enclosures are suitable for studying swarming and mating in laboratory-bred males in field-like settings, providing an important reference for future studies aimed at assessing the comparative mating ability of strains for SIT and other vector control strategies.