Ejaculate Ejection by Female Fruit Flies Does Not Correlate With Mating Latency or Male Presence, Irrespective of How Ejection Is Measured

Sexual selection operates across pre- and post-mating episodes, driven by intra-sexual competition and inter-sexual choice for mates or gametes. While these episodes can act in the same direction, this is not always the case; for example, males who are preferred as mates by females may produce sperm not preferred for fertilisation. In many species, post-mating sexual selection is measured using ejaculate ejection behaviour. However, whether different proxies for this behaviour correlate with each other, and whether the environment experienced by females post-mating influences ejaculate ejection, has received little attention until recently. Using <i>Drosophila melanogaster</i>, we first explore whether mating latency, a proxy for inter-sexual pre-mating preference, correlates with different proxies for female post-mating preference (ejection latency, ejection likelihood, numbers of sperm ejected, numbers of sperm retained and proportion of sperm ejected). We further test whether the presence of a male in the female's post-mating environment influences such ejaculate ejection behaviour. We find no effect of male presence. Additionally, mating latency does not correlate with any measure of ejaculate ejection behaviour, possibly indicating the decoupling of sexual selection at pre- and post-mating stages. Importantly, we find that ejection latency, commonly used to quantify female post-mating preference, does not correlate with the numbers of sperm ejected or retained, or the proportion of sperm ejected. Instead, females that eject more sperm also retain more sperm due to variation in sperm numbers inseminated by males. This highlights the importance of using different proxies for ejaculate ejection behaviour in tandem and carefully interpreting their biological meaning.