We examined the systematics of round-eared bats (Tonatia and Lophostoma) using sequence data from the nuclear recombination activator gene-2 (Rag2) gene and the mitochondrial valine transfer RNA, 12S ribosomal RNA (rRNA), and 16S rRNA. Some analyses of the mitochondrial and combined data sets indicate that round-eared bats may be paraphyletic relative to the genera Phyllostomus, Phylloderma, and Mimon. Both nuclear and mitochondrial data sets indicate substantial genetic divergence between Tonatia and Lophostoma, and neither data set unambiguously demonstrates monophyly of round-eared bats. However, our analyses cannot exclude the possibility that round-eared bats form a monophyletic lineage that separated anciently into Tonatia and Lophostoma. Within the Tonatia lineage, T. bidens and T. saurophila are sister taxa but are divergent for both nuclear and mitochondrial sequences. Nuclear data suggest that L. silvicolum may be paraphyletic, as currently recognized, because L. evotis appears within this clade.