The widespread family Vespertilionidae constitutes the largest family of bats with 533 described species. However, systematic relationships within this family remain unresolved for many clades among the pipistrelle-like bats of the tribes Vespertilionini and Pipistrellini. In this study, we focus on the recently proposed endemic African subtribe Laephotina (Vespertilionini) and African representatives of the Pipistrellini. Recent revisionary systematics utilizing mitochondrial, morphological, and morphometric data has clarified relationships and species limits in Laephotina (Laephotis, Neoromicia, Afronycteris, Pseudoromicia), correcting genus assignments and describing two new genera. However, genetic data from independent nuclear loci have been lacking to date. Using four independent nuclear introns to generate gene trees and species trees, we establish strong support for the monophyly of the four Laephotina genera and the sister relationship of Laephotis + Neoromicia. However, the relationships of Afronycteris and Pseudoromicia remain unresolved. Our results are consistent with mitochondrial, karyotypic, and bacular morphological data previously used to delimit Laephotina. We found strong support for Nycticeinops + Afropipistrellus as sister to Laephotina. Deep divergence between Nycticeinops and Afropipistrellus in nuclear introns adds to dental and bacular evidence for their recognition as separate genera.