Reproductive output of old polygynous males is limited by seminal fluid, not sperm number

Abstract

Advancing male age can lead to reproductive senescence, which in males is thought to be largely driven by declines in the numbers of sperm transferred by old males. This decline is predicted to be particularly pronounced in polygynous species, where males become sperm limited over a mating sequence. However, males also transfer seminal fluid to females, and little is known about the contribution of seminal fluid to constrain the reproductive output of old, multiply-mating males. Using Drosophila melanogaster, we investigated whether age-related variation in male reproductive output is driven by differential limitation of sperm or seminal fluid, over a series of experimental matings. Consistent with reproductive senescence, old males produced fewer offspring than young males. However, this pattern was not driven by sperm limitation, with old males having more sperm and transferring similar numbers of sperm to a female, compared to young males. Yet surprisingly, females stored fewer sperm when mated to old than young males. Notably, females mated to old multiply-mating males produced more offspring when supplemented with seminal fluid, suggesting that fertility of old males over successive matings was limited by seminal fluid availability. Generally, our study indicates that germline maintenance might be prioritised over somatic maintenance as hypothesized by the disposable soma theory of ageing, and that seminal fluid senescence is a key contributor of reproductive decline with age. While other factors such as differential sperm viability and female post-mating responses could have also influenced our results, our study highlights the under-appreciated role of seminal fluid in mediating male reproductive senescence in polygynous species. Significance statementA key assumption in ageing research is that old males are less fertile than young males, and that this reduced fertility is partly driven by old males producing fewer sperm. However, senescence in male fertility can be caused via other ejaculate-mediated pathways, which we investigate using fruit flies. Contrary to expectations, we reveal that senescence in male fertility is not because of declines in male sperm reserves, but is instead due to age-related changes in seminal fluid and differences in female sperm storage with male age. These declines in the reproductive output of old, multiply-mating males are alleviated by supplementing females with "extra" seminal fluid. Our study demonstrates that male reproductive senescence is reversible, highlighting the underappreciated role of seminal fluid in modulating senescence in polygynous species. These results have potential for improving animal fertility and our understanding of sexual selection.