New stands of Paramecium tredecaurelia, a rare species of the P. aurelia spp. complex, were identified in Thailand and Madagascar on the basis of mating reactions and molecular markers (rDNA and mtDNA). Analysis of DNA fragments showed that all P. tredecaurelia strains, the recently recorded ones and the ones known previously from France, Mexico, and Israel, form a monophyletic and well-defined clade in the P. aurelia species trees. All of these strains, collected from different localities around the world, represent identical or nearly identical haplotypes in terms of all the studied DNA fragments. Given the huge distances between particular collection sites, such a low level of variability of the studied sequences may result from a slow rate of evolution in P. tredecaurelia.