The male mosquito contribution towards malaria transmission: Mating influences the Anopheles female midgut transcriptome and increases female susceptibility to human malaria parasites

Abstract

Mating causes dramatic changes in female physiology, behaviour, and immunity in many insects, inducing oogenesis, oviposition, and refractoriness to further mating. Females from the Anopheles gambiae species complex typically mate only once in their lifetime during which they receive sperm and seminal fluid proteins as well as a mating plug that contains the steroid hormone 20-hydroxyecdysone. This hormone, which is also induced by blood-feeding, plays a major role in activating vitellogenesis for egg production. Here we show that female Anopheles coluzzii susceptibility to Plasmodium falciparum infection is significantly higher in mated females compared to virgins. We also find that mating status has a major impact on the midgut transcriptome, detectable only under sugar-fed conditions: once females have blood-fed, the transcriptional changes that are induced by mating are likely masked by the widespread effects of blood-feeding on gene expression. To determine whether increased susceptibility to parasites could be driven by the additional 20E that mated females receive from males, we mimicked mating by injecting virgin females with 20E, finding that these females are significantly more susceptible to human malaria parasites than virgin females injected with the control 20E carrier. Further RNAseq was carried out to examine whether the genes that change upon 20E injection in the midgut are similar to those that change upon mating. We find that 79 midgut-expressed genes are regulated in common by both mating and 20E, and 96% (n = 76) of these are regulated in the same direction (up vs down in 20E/mated). Together, these findings show that male Anopheles mosquitoes induce changes in the female midgut that can affect female susceptibility to P. falciparum. This implies that in nature, males might contribute to malaria transmission in previously unappreciated ways, and that vector control strategies that target males may have additional benefits towards reducing transmission.

Author summary

Malaria mosquitoes must successfully mate and bloodfeed in order to reproduce. The impact of bloodfeeding on malaria transmission is clear given that all transmission is caused by female mosquitoes that have fed at least twice: once leading to an initial infection, and again 10–14 days later resulting in parasite transmission. The impact of mating on malaria transmission is less clear. Here we show that mating status significantly enhances transmission, such that mated females are more likely to transmit malaria parasites than virgin females. We further examine whether a hormone transferred by mating might cause this enhanced susceptibility, and we find that indeed the receipt of this hormone is also correlated with enhanced susceptibility. The results of this study imply that efforts to target male mosquitoes might not only suppress mosquito populations, but also act to decrease vector competence among residual females.