The Callitrichaceae are a monogeneric family of aquatic angiosperms comprising approximately 50 terrestrial, amphibious, and obligately submersed species. Callitriche is unique in being the only known genus with co-occurring aerial and underwater pollination systems. Mature pollen structure is correlated with growth habit, pollination biology, and phylogeny within the genus. In the present study, development of exineless pollen in the obligately submersed species Callitriche truncata was examined, with particular emphasis on the tetrad stage. Pollen ontogeny occurred rapidly and non-synchronously; tetrads, free microspores, and two-celled pollen grains were identified within the same anthers. Formation of the intine also occurred relatively early, during the tetrad stage. Tetrads were surrounded by a structurally distinct envelope, and its ultrastructure and histochemistry indicate that this callose-like envelope is in a transitional state. Reduction or complete loss of the exine has evolved at least twice in Callitrichaceae, and the new ontogenetic data indicate that exine loss evolves more quickly than the loss of callose. In addition, developmental information on exineless pollen in C. truncata coupled with other palynological data for the exine-bearing terrestrial and amphibious growth forms provide support for the hypothesis that underwater pollination has had a relatively recent origin in the family.