Mechanisms of sperm competition revealed by sperm viability assessment using live/dead dual fluorescence

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Female damselflies in the family Calopterygidae have two sperm-storage organs, a spherical bursa copulatrix and a tubular spermatheca; males possess a peculiar aedeagus with a recurved head, with which to remove bursal sperm, and lateral spiny processes to remove spermathecal sperm. The lateral processes, however, differ among species or populations in terms of their width relative to the spermathecal duct; the narrower processes are physically able to access spermathecal sperm, while the wider ones are not. In this study, sperm-storage patterns and aedeagal structures were compared between two calopterygid species with different spermathecal structures, Calopteryx cornelia and Mnais pruinosa, with respect not only to sperm quantity (number) but also to sperm quality (viability), by using a recently developed method based on live/dead dual fluorescence. Calopteryx cornelia is a typical spermathecal sperm remover. In this species, viability was similar between bursal and spermathecal sperm. On the other hand, in M. pruinosa the spermatheca was much smaller than the bursa and often contained no sperm. Even when the spermatheca of this species did contain sperm, a high percentage was dead. While the spermatheca of M. pruinosa has such atrophic tendencies, males, have developed long and spiny lateral processes similar to those of C. cornelia. This suggests functions other than spermathecal sperm removal. The lateral processes possibly function as stoppers or guides for manipulating the aedeagal head to remove the sperm mass from the bursa.