Microsatellite analysis of sperm-use patterns in the bushcricket Requena verticalis

The proportion of offspring sired by the second male to mate with a doubly mated female, Pt, is a ubiquitously measured statistic in the study of inse ct sperm competition. Nevertheless, the underlying mechanisms of sperm tran sfer, storage, and use that determine Pt are poorly understood. Typically t he second male to mate gains moderate to high paternity. More rarely, the f irst male to mate gains the majority of fertilizations. Here we examine the transfer, storage, and use of sperm in the bushcricket Requena verticalis, a species with male parental investment and almost complete first male pat ernity. Sperm drain from an externally attached spermatophore into the fema le's reproductive tract, where they are transported to the sperm store or s permatheca. We find that only sperm from the first male to mate are transpo rted to the spermatheca. We provide some data that address a number of diff erent mechanisms that might account for the lack of storage of second-male sperm. DNA microsatellite markers are developed to assign paternity. By man ipulating the numbers of sperm transferred by first and second males, we sh ow that the size of the ejaculate transferred by the first male has a major impact on paternity; increasing ejaculate size of the first male assures h is paternity. Paternity assurance in R. verticalis holds significant implic ations for the evolution of paternal investment via the male's nuptial gift .