The foliar epidermis and cuticle ofStaurogyne longifolia(Nees) Kuntze,Elytraria acaulis(L.f.) Lindau var.acaulis, E. acaulisvar.lyrata(Nees) Bremek. andNelsonia campestrisR.Br, have been investigated, revealing broad similarities with those of the rest of the Acanthaceae; the presence of diacytic stomata in the Nelsonioideae is evidence of its affinity with the Acanthaceae in general‐, while the presence of panduriform glandular hairs and the absence of the cystoliths in Nelsonioideae indicate its particular affinity with the Thunbergioideae. Substantial evidence is provided to support the retention of Nelsonioideae as a subfamily of the Acanthaceae, rather than its transfer to the Scrophulariacea