Cryptic female choice in response to male pheromones in Drosophila melanogaster

Females control the paternity of their offspring by selectively mating with males they perceive to be of high quality. In species where females mate with multiple males in succession, females may bias offspring paternity by favoring the sperm of one male over another, a process known as cryptic female choice (CFC). While evidence of CFC exists in multiple taxa, the mechanisms underlying this process have remained difficult to unravel. Understanding CFC requires demonstration of a female-driven post mating bias in sperm use and paternity, and a causal link between this bias and male cues. Here, we show that in the vinegar fly Drosophila melanogaster, mated females eject the ejaculate of their first mate faster when exposed to the pheromones of an attractive male than in the presence of an unattractive one. Using transgenic males expressing fluorescent sperm, we show that exposure to attractive males between mating causes twice-mated females to bias sperm storage towards the second male, affecting paternity. Using pheromonal bioassays in combination with genetic manipulation of sensory systems, we show that females modulate ejaculate ejection latency in response to male pheromones heptanal and 11-cis-Vaccenyl acetate (cVA) sensed via olfactory receptor neurons OR35a, Or22a, Or65a and OR67d, demonstrating that polyandrous females use male pheromonal cues to modulate ejaculate ejection timing. We provide the first demonstration to our knowledge of a CFC mechanism allowing a female to increase or decrease the share of paternity of her first mate depending on the sensing of the quality of potential mates in her environment. These findings showcase that paternity can be influenced by events that go beyond copulation and highlights the importance of post-copulatory sexual selection. One Sentence SummaryWe show that females bias sperm use and paternity towards specific males in response to male pheromones.