Sperm ultrastructure and the number of sperm per bundle were studied in twoDrosophilaspecies from the tripunctata radiation,D. dunniandD. cardini.The number of sperm per bundle in both species was found to have an unusual frequency distribution, as none of the bundles obeyed the 4(2”) rule proposed for species with definitive spermatogonia division. The absolute and relative size of the cross sectional areas of the mitochondrial derivatives ofD. dunniandD. cardinisperm was found to be somewhat different fromD. hydei(repleta radiation), and very different fromD. melanogasterof the Sophophora radiation. Lastly, a mass of residual material and a desmosome-like attachment plaque were demonstrated between the axoneme and the large mitochondrial derivative inD. dunniandD. cardinisperm. This study highlights the importance of ultrastructural studies of spermatogenesis in the analysis of the phylogeny ofDrosophila.