Sperm structure and locations in the hermaphroditic freshwater snail <i>Planorbella trivolvis</i>

ABSTRACT The freshwater snail Planorbella trivolvis provides a model system for investigating hermaphrodite reproduction. Self-fertilization is rare, and individuals may mate as male, female, or reciprocally; after mating, sperm is stored with recipients laying eggs for 16–18 weeks after a single mating. Some key unanswered questions in this and other related species involve sperm. Very little is known about sperm structure, the pathway of sperm post-copulation or the location of sperm in the hours just after mating or in the long term. We carried out experiments to describe and determine the location of sperm produced by snails (autosperm) as well as that of sperm received after copulation (allosperm) using staining and fluorescence microscopy. We searched for and described sperm using phase contrast microscopy and used the DNA stain DAPI to visualize their nuclei with fluorescence microscopy. Sperm was found primarily in the seminal vesicles but also in the ovotestes and hermaphroditic duct; these cells have elongate cone-shaped heads with a helical keel and long helically twisted tails. We then performed mating experiments to track sperm location post-copulation. We incubated snails in a treated tap water with the less toxic stain Hoechst 33342 to label sperm in donor snails, mated them to unstained individuals, then tracked the location of sperm in recipients either several days after mating or weeks later. Just after mating, sperm was primarily located in the sperm receptacle sac, while long-term storage appeared to occur in or near the seminal vesicles and hermaphroditic duct. Further analysis will be necessary to determine how sperm are allocated for fertilization, particularly how autosperm are distinguished from allosperm.