Ancient asexual animal groups, such as bdelloid rotifers and darwinuloid ostracods, are excellent model organisms to study the effects of long-term asexuality. However, the absolute length of time that these groups have been fully asexual is mostly ignored. In the case of the darwinuloid ostracods, the fossil record shows that sexual reproduction disappeared almost completely after the end of Permian mass extinction (ca. 245 Myr ago), although several putative records of males from the Mesozoic obscure the exact time-frame of obligate asexuality in darwinuloids. Here, we re-examine the Mesozoic darwinuloid records, with regard to the reproductive mode of the assemblages. Three criteria to distinguish males in fossil populations (lack of brood pouch, position of muscle scars and size dimorphism) are used here to test for the presence of males in darwinuloid assemblages. A large, well-preserved assemblage of Darwinula leguminella (Forbes 1885) from the latest Jurassic (ca. 145 Myr ago) of England is found to be markedly variable in size and shape, but nevertheless turns out to be an all female assemblage. The exceptional preservation of the material also allows the re-assignment of this species to the extant darwinuloid genus Alicenula. All other putative dimorphic darwinuloid records from the Mesozoic are re-examined using the same criteria. The hypothesis that these assemblages represent bisexual populations is rejected for all post-Triassic (ca. 208 Myr ago) records.