The increase of DNA sequence data in public data bases makes it possible to distinguish species using a quantitative method based on measurements of genetic distances.We used two monophyletic genera Cetrelia and Platismatia of Cetrarioid lichens (Ascomycota) as the examples.The new nucleotide substitution model was developed and the software GenCAT (Genetic Coherence Analysis Tool) was modified for calculating the genetic distances.Intraspecific and interspecific distances inferred from rDNA ITS sequences were examined.Twenty-six samples for 5 species with 15 haplotypes in Cetrelia and 24 samples for 6 species with 15 haplotypes in Platismatia were analyzed.The genetic distances within and among species were well separated in most cases allowing the calculation of a threshold for species delimitation, with exception in the case ofPlatismatia herrei and P.stenophylla, which may belong to the same species with a Pacific coastal distribution in North America.