The ciliate genus Spirostomum comprises eight morphospecies, inhabiting diverse aquatic environments worldwide, where they can be used as water quality indicators. Although Spirostomum species are relatively easily identified using morphological methods, the previous nuclear rDNA-based phylogenies indicated several conflicts in morphospecies delineation. Moreover, the single locus phylogenies and previous analytical approaches could not unambiguously resolve phylogenetic relationships among Spirostomum morphospecies. Here, we attempt to investigate species boundaries and evolutionary history of Spirostomum taxa, using 166 new sequences from multiple populations employing one mitochondrial locus (CO1 gene) and two nuclear loci (rRNA operon and alpha-tubulin gene). In accordance with previous studies, relationships among the eight Spirostomum morphospecies were poorly supported statistically in individual gene trees. To overcome this problem, we utilised for the first time in ciliates the Bayesian coalescent approach, which accounts for ancestral polymorphisms, incomplete lineage sorting, and recombination. This strategy enabled us to robustly resolve deep relationships between Spirostomum species and to support the hypothesis that taxa with compact macronucleus and taxa with moniliform macronucleus each form a distinct lineage. Bayesian coalescent-based delimitation analyses strongly statistically supported the traditional morphospecies concept but also indicated that there are two S. minus-like cryptic species and S. teres is non-monophyletic. Spirostomum teres was very likely defined by a set of ancestral features of lineages that also gave rise to S. yagiui and S. dharwarensis. However, molecular data from type populations of the morphospecies S. minus and S. teres are required to unambiguously resolve the taxonomic problems.

Pollution after oil spill represents extreme habitat for survival and is a major concern for loss of species diversity in the affected area. In this study, we investigated soil samples collected from a petrochemical industry, Ulsan, South Korea. The soil was in the phase of recovery from the contamination of crude oil spill. Detailed investigation, based on morphology, ontogenesis, and molecular phylogenetic methods, resulted in discovery of a novel hypotrich ciliate, i.e., Metasterkiella koreana n. gen., n. sp., which is morphologically characterized by a semirigid body, undulating membranes in Oxytricha pattern, 18 frontal-ventral-transverse cirri with cirrus V/3 placed posteriorly, one right and one left row of marginal cirri, four dorsal kineties, two dorsomarginal rows, and caudal cirri at the end of dorsal kineties 1, 2, and 4. Interestingly, during ontogenesis, formation of three common anlagen for the proter and the opisthe and involvement of cirrus V/3 in anlagen formation was observed. The dorsal ontogenesis was typical of oxytrichids, i.e., simple fragmentation of dorsal kinety 3 and formation of dorsomarginal rows close to the right marginal row. The new species was found to be similar with Sterkiella subtropica, except for some minor differences in morphometry, and at gene level with only one base pair difference. In phylogenetic analyses, based on SSU rRNA gene sequence, M. koreana cluster in a clade away from Sterkiella species, which could be explained by the differences in the morphogenetic pattern between these two genera. It is proposed that S. subtropica probably belongs to Metasterkiella; however, we do not perform changes and wait for the reinvestigation of its morphogenetic pattern.