Comments on species divergence in the genus Sphaerium (Bivalvia) and phylogenetic affinities of Sphaerium nucleus and S. corneum var. mamillanum based on karyotypes and sequences of 16S and ITS1 rDNA.

Chromosome, 16S and ITS1 rDNA sequence analyses were used to obtain reliable diagnostic characters and to clarify phylogenetic relationships of sphaeriid bivalves of the genus Sphaerium. The species studied were found to be diploid, with modal number 2n = 28 in S. nucleus and 2n = 30 in S. corneum var. mamillanum. Small, biarmed, C- negative B chromosomes were found in all studied populations of both species. Karyological and molecular markers revealed no differences between S. corneum s. str. and S. corneum var. mamillanum. No intraspecific differences were found in the basic karyotype of S. nucleus. Molecular analyses, however, uncovered three genetically distinct ITS1 lineages: one comprised of samples from Lithuania, Slovakia, and Russia, another from Czech, and a third from Ukraine. Additionally to known 16S haplotype from Ukraine, three new 16S haplotypes of S. nucleus were detected: one in the samples from Lithuania and Russia, one in Slovakian and one in Czech population. In the ITS1 phylogenetic tree, all branches of S. nucleus clustered in one clade. In the 16S phylogenetic tree, however, the haplotype of Czech S. nucleus formed a separate branch, distant from three other haplotypes of S. nucleus. Molecular results indicate that in the context of the Evolutionary Species Concept the S. nucleus morphospecies may represent a complex of separate taxa, however referring on the Biological Species Concept the genetic lineages could represent the intraspecific variability.