Abstract Background In this study, we investigate species limits in the cyanobacterial lichen genus Rostania (Collemataceae, Peltigerales, Lecanoromycetes). Four molecular markers (mtSSU rDNA, β-tubulin, MCM7, RPB2) were sequenced and analysed with two coalescent-based species delimitation methods: the Generalized Mixed Yule Coalescent model (GMYC) and a Bayesian species delimitation method (BPP) using a multispecies coalescence model (MSC), the latter with or without an a priori defined guide tree. Results Species delimitation analyses indicate the presence of eight strongly supported candidate species. Conclusive correlation between morphological/ecological characters and genetic delimitation could be found for six of these. Of the two additional candidate species, one is represented by a single sterile specimen and the other currently lacks morphological or ecological supporting evidence. Conclusions We conclude that Rostania includes a minimum of six species: R. ceranisca, R. multipunctata, R. occultata 1, R. occultata 2, R. occultata 3, and R. occultata 4,5,6. Three distinct Nostoc morphotypes occur in Rostania, and there is substantial correlation between these morphotypes and Rostania thallus morphology.