The seminal odorant binding protein Obp56g is required for mating plug formation and male fertility in .

In and other insects, the seminal fluid proteins (SFPs) and male sex pheromones that enter the female with sperm during mating are essential for fertility and induce profound post-mating effects on female physiology and behavior. The SFPs in and other taxa include several members of the large gene family known as odorant binding proteins (Obps). Previous work in has shown that some genes are highly expressed in the antennae and can mediate behavioral responses to odorants, potentially by binding and carrying these molecules to odorant receptors. These observations have led to the hypothesis that the seminal Obps might act as molecular carriers for pheromones or other compounds important for male fertility in the ejaculate, though functional evidence in any species is lacking. Here, we used RNAi and CRISPR/Cas9 generated mutants to test the role of the seven seminal Obps in fertility and the post-mating response (PMR). We found that is required for male fertility and the induction of the PMR, whereas the other six genes had no effect on fertility when mutated individually. is expressed in the male's ejaculatory bulb, an important tissue in the reproductive tract that synthesizes components of the mating plug. We found males lacking fail to form a mating plug in the mated female's reproductive tract, leading to ejaculate loss and reduced sperm storage. We also examined the evolutionary history of these seminal genes, as several studies have documented rapid evolution and turnover of SFP genes across taxa. We found extensive lability in gene copy number and evidence of positive selection acting on two genes, and . Comparative RNAseq data from the male reproductive tract of multiple species revealed that shows high male reproductive tract expression only in species of the and groups, though conserved head expression in all species tested. Together, these functional and expression data suggest that may have been co-opted for a reproductive function over evolutionary time.