Copulation behavior has often been shaped by sexually selected sperm competition or cryptic female choice. However, manipulation of previously deposited ejaculates is unknown in the social Hymenoptera and the degree to which sperm competes after insemination or is actively selected by females has remained ambiguous. We studied the mating process in the leaf-cutting ants Atta colombica and A. cephalotes, which belong to one of the few derived social insect lineages where obligate multiple mating has evolved. As copulations often occur at night and in remote places, direct observations were impossible, so we had to reconstruct the sequential copulation events by morphological analysis of the male and female genitalia and by tracking the process of sperm transfer and sperm storage. We show that Atta male genitalia have two external rows of spiny teeth, which fit into a specialized pouch organ in the female sexual tract. Reconstruction of the sperm storage process indicated that sperm is transferred to the spermatheca during or immediately after ejaculation and without being mixed with sperm and seminal fluids from other males. A convergent mechanism of direct sperm transfer to the spermatheca of queens is known from two species of dwarf honeybees. Direct sperm transfer may restrict female control over the sperm storage process and the number of males that contribute to the stored sperm.
Keywords: Animals; Ants; Female; Flight, Animal; Genitalia, Female; Genitalia, Male; Male; Plant Leaves; Sexual Behavior, Animal; Spermatozoa