Abstract Background Freshwater ecosystems are among the most endangered habitats on Earth, with approximately one-fourth of aquatic species at risk of extinction. Effective conservation efforts require comprehensive monitoring and accurate species identification, including often overlooked groups. Planarian flatworms are one such group that, although commonly present in freshwater ecosystems worldwide, remains understudied even in species-rich areas, e.g. Croatia. As a result, the true extent of planarian diversity often remains underappreciated. Results With the goal of characterising the Croatian planarian diversity, we used an integrative approach combining barcoding and classical taxonomy methods. Motivated by the highly skewed representation of planarian diversity in current GenBank records of the barcoding gene cytochrome c oxidase subunit I (COI), we first optimised primer design and amplification protocols. Applying these approaches to field-collected material from Croatia, we substantially expanded the number of taxonomically curated COI barcode sequences for European dugesiids, dendrocoelids and planariids. In addition, our efforts resulted in the description of a new pigmented Dendrocoelum species, Dendrocoelum pigmentatum Vila-Farré, sp. nov. , the discovery of two highly differentiated haplotypic clades in Schmidtea lugubris , and the rediscovery of Polycladodes alba in Croatia after a century. Conclusions Overall, our effort integrates Croatia as an underexplored but planaria species-rich region into the endeavour to systematically describe the planarian fauna of Europe. The expansion of a known number of Croatian planarian species from eight to sixteen and the discovery of a new, large planarian species in continental Europe, Dendrocoelum pigmentatum , demonstrate the effectiveness of our integrative approach. Overall, our work highlights the underappreciated diversity of planarians, even in continental Europe and supports practical conservation efforts to preserve aquatic biodiversity.
Vila-Farré et al. (Mon,) studied this question.