Self-fertilization, sex allocation and spermatogenesis kinetics in the hypodermically inseminating flatworm Macrostomum pusillum .

The free-living flatworm genus Macrostomum is an emerging model system for studying the links between sex allocation, sexual selection and mating system evolution, as well as the underlying developmental and physiological mechanisms responsible for wide intra- and inter-specific variability in reproductive phenotypes. Despite compelling comparative morphological evidence of sexual diversity, detailed experimental work on reproductive behaviour and physiology in Macrostomum has so far been largely limited to just two species, M. lignano and M. hystrix , an obligate and a preferential outcrosser, respectively. In this study, we establish that a third species, M. pusillum , exhibits a combination of reproductive traits strikingly different from both of its congeners. Unlike M. lignano , we demonstrate that M. pusillum does not adjust sex allocation or the speed of spermatogenesis to the prevailing social group size. M acrostomum pusillum 's relatively simple sperm morphology likely explains the short spermatogenesis duration we report, and is linked to a hypodermically inseminating mode of fertilization, which we show also means that these worms are capable of self-fertilization. Surprisingly, and unlike M. hystrix , selfing in isolated worms commences after only a short (if any) delay compared with the onset of reproduction in grouped individuals, with little evidence of differential inbreeding depression in 'isolated' progeny. These combined results suggest that, in nature, M. pusillum may be regularly selfing, in contrast to the congeners studied to date. Our findings highlight the rapid and correlated evolution of reproductive traits, and reinforce the utility of the genus Macrostomum for understanding the evolutionary and developmental mechanisms responsible for this diversity.