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ABSTRACT Freshwater planarians constitute an important component in aquatic ecosystems as predators. They are, nonetheless, delicate animals used as indicators of water quality. This group has been little studied in The Antilles, where only seven species of Girardia Ball, 1974 have been reported. Those records date from the last two centuries and were identified based on morphology, leaving several specimens unidentified. Furthermore, the anatomical similarities among species and the lack of the copulatory apparatus in fissiparous populations make it necessary to use molecular data to perform accurate species delimitations and phylogenetic studies. The Cuban archipelago is the reservoir of the highest species diversity in the Caribbean. However, only one species of freshwater triclad has been described, Girardia cubana (Codreanu & Balcesco, 1973), which is endemic to Cuba. Recent samplings in the western part of the island molecularly identified Girardia sinensis Chen & Wang, 2015. At present, we are performing broad samplings all around Cuba. As a first result, we here present a phylogeny-based identification of freshwater planarians, collected in four localities of eastern Cuba, inferred using nuclear and mitochondrial markers. The presence of G. sinensis in the eastern part of the island is reported and two other lineages of the genus are identified, at least one could be a new species. Moreover, we found a lineage belonging to Cavernicola, of which there are no previous records in The Antilles. These findings support that the planarian richness of Cuba has been underestimated and new species could be described, providing relevant biogeographic information about the group in the Caribbean.

Freshwater planarians of the genus Girardia have been introduced all over the world, but little is known about the species involved and their possible impact on autochthonous ecosystems. Using molecular phylogenetics and niche modelling under different climatic scenarios we examine the human-induced spread of alien Girardia species from their original areas of distribution in the Americas to other areas. Our results corroborate that Girardia populations spreading worldwide belong to three species: G. dorotocephala, G. sinensis, and G. tigrina. Our study emphasizes that G. sinensis is native to North America and shows that G. dorotocephala has a broader range of introduced localities than previously known. Niche modelling revealed that the three species have a broad range of potential distribution in extensive regions of the Northern Hemisphere. Regardless of the future climatic scenario, their distributional range will increase towards northern Europe, without diminishing the high suitability of regions in the south. Their environmental requirements, being generalists with high suitability for human-modified habitats, and fissiparous reproduction explain their successful colonization. In the Iberian Peninsula, G. tigrina and G. sinensis have extensive areas of high suitability, overlapping with the more limited suitable areas of autochthonous planarians, pointing to potential detrimental effects of Girardia invaders.

The application of road de-icing salts has the potential to salinize fresh waters and degrade habitat for aquatic organisms. In the Australian Alps, the ecological effects of even small salinity increases from de-icing may be different than in North America and Europe because of (1) differences in the evolutionary history, and (2) areas with de-icing in Australia are not located in urbanized landscapes where de-icing has been largely studied elsewhere. In this study, we tried to determine the salinity increases attributable to de-icing in Australia and the effects of this increase in salinity to stream macroinvertebrates. We observed increased salt concentrations (as measured by continuous measurements of electrical conductivity (EC) and periodic measurements of chloride concentrations) in streams near two Australian ski resorts, during the snow seasons (June to September) of 2016 to 2018. The maximum EC observed in streams in salted sites near Perisher, New South Wales, was 390 µS cm−1 compared with a maximum of 26.5 µS cm−1 at unsalted sites. Lower EC values (i.e., maximum 61.1 µS cm−1) and short durations of salinity increases in streams near Falls Creek, Victoria, were not expected to cause an adverse biological response. Salt storage in the landscape was evident at salted sites near Perisher where EC was above background levels during periods of the year when no salt was applied to roads. Stream macroinvertebrate community composition differed at sites receiving run-off from road salting activities near Perisher. Abundances of Oligochaeta (worms) (up to 11-fold), Dugesiidae (flat worms) (up to fourfold), and Aphroteniinae (chironomids) (up to 14-fold) increased, whereas Leptophlebiidae (mayflies) decreased by up to 100% compared with non-salted sites. The taxa that were less abundant where de-icing salts were present tended to be the same taxa that toxicity testing revealed to be relatively salt sensitive species. This study demonstrates a causal link between de-icing salts, elevated stream salinity, and altered macroinvertebrate community composition in streams that received run-off from road de-icing activity in the Australian Alps.

Two new species of the genus Dugesia from Hainan Island and Leizhou Peninsula are described by applying an integrative approach, including morphological, karyological, histological, and molecular information. In the molecular phylogenetic tree, the two new species, Dugesia saccata Chen & Dong, sp. nov. and Dugesia aconcinna Chen & Dong, sp. nov. , fall into an Eastern Palearctic/Oriental clade and an Oriental/Australasian clade, respectively, while sharing only a rather distant relationship. The separate specific status of the two new species is supported also by their genetic distances. Dugesia saccata is characterised by the presence of the following features: a sac-shaped expansion at the knee-shaped bend of the bursal canal; ventrally displaced ejaculatory duct with a subterminal opening; a duct between diaphragm and seminal vesicle; mixoploid karyotype with diploid complements of 2n = 2x = 16 and triploid complements of 2n = 3x = 24, with all chromosomes being metacentric. Dugesia aconcinna is characterised by the presence of the following features: asymmetrical openings of the oviducts into the bursal canal and the common atrium, with the left oviduct opening into the common atrium and the right oviduct opening into the most ventral, proximal portion of the bursal canal, at the point where the latter communicates with the common atrium; vasa deferentia separately opening into the posterior portion of the seminal vesicle; penis papilla of a very characteristic shape, with the part housing the connecting duct, diaphragm, and ejaculatory duct being a cylindrical structure with a blunt tip, while at its right-hand side sits a large penial fold that attaches to the base of the penis papilla; ejaculatory duct following a ventrally displaced course through the penis papilla, after which it opens at the tip of the papilla; presence of a duct between diaphragm and seminal vesicle.

Mitochondrial genomes (mitogenomes) are useful and relatively accessible sources of molecular data to explore and understand the evolutionary history and relationships of eukaryotic organisms across diverse taxonomic levels. The availability of complete mitogenomes from Platyhelminthes is limited; of the 40 or so published most are from parasitic flatworms (Neodermata). Here, we present the mitogenomes of two free-living flatworms (Tricladida): the complete genome of the freshwater species Crenobia alpina (Planariidae) and a nearly complete genome of the land planarian Obama sp. (Geoplanidae). Moreover, we have reanotated the published mitogenome of the species Dugesia japonica (Dugesiidae). This contribution almost doubles the total number of mtDNAs published for Tricladida, a species-rich group including model organisms and economically important invasive species. We took the opportunity to conduct comparative mitogenomic analyses between available free-living and selected parasitic flatworms in order to gain insights into the putative effect of life cycle on nucleotide composition through mutation and natural selection. Unexpectedly, we did not find any molecular hallmark of a selective relaxation in mitogenomes of parasitic flatworms; on the contrary, three out of the four studied free-living triclad mitogenomes exhibit higher A+T content and selective relaxation levels. Additionally, we provide new and valuable molecular data to develop markers for future phylogenetic studies on planariids and geoplanids.

Two new species of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Hainan Island of China are described on the basis of morphological, karyological and molecular data. Dugesia semiglobosa Chen & Dong, sp. nov . is mainly characterized by a hemispherical, asymmetrical penis papilla with ventrally displaced ejaculatory duct opening terminally at tip of penis papilla; vasa deferentia separately opening into mid-dorsal portion of intrabulbar seminal vesicle; two diaphragms in the ejaculatory duct; copulatory bursa formed by expansion of bursal canal, lined with complex stratified epithelium, which projects through opening in bursa towards intestine, without having open communication with the gut; mixoploid chromosome complement diploid (2n = 16) and triploid (3n = 24), with metacentric chromosomes. Dugesia majuscula Chen & Dong, sp. nov . is mainly characterized by oviducts opening asymmetrically into female reproductive system; hyperplasic ovaries; expanded posterior section of bursal canal; vasa deferentia separately opening into mid-dorsal portion of seminal vesicle; asymmetrical penis papilla due to ventral course of ejaculatory duct, which has subterminal and dorsal opening at tip papilla; mixoploid chromosome complement diploid (2n = 16) and triploid (3n = 24); chromosomes metacentric. Apart from their anatomy, separate species status of the two new species is supported also by their genetic distances and by their positions in the phylogenetic tree. The sexualization process may have been induced by the lower temperatures, in comparison with their natural habitat, under which the worms were cultured in the laboratory.

Two new species of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from the tropical monsoon forest in southern China are described on the basis of an integrative taxonomic study involving morphology, karyology, histology, and molecular analyses. The new species Dugesia circumcisa Chen & Dong, sp. nov. is characterised by asymmetrical openings of the oviducts; right vas deferens opening at anterior portion of the seminal vesicle and the left one opening at mid-lateral portion of the seminal vesicle; two diaphragms in ejaculatory duct, the latter being ventrally displaced and opening at the tip of the penis papilla, which is provided with a nozzle; wide duct connecting male atrium and common atrium; chromosome complement triploid with 24 metacentric chromosomes. The other new species, Dugesia verrucula Chen & Dong, sp. nov. , is characterised by the large size of the living worm, usually exceeding 3.5 cm in length; asymmetrical openings of the oviducts; subterminal opening of ventrally displaced ejaculatory duct; vasa deferentia symmetrically opening into the postero-lateral portion of the seminal vesicle; well-developed duct between the seminal vesicle and diaphragm; single dorsal bump near the root of the penis papilla; bursal canal with pleated wall and spacious posterior section; unstalked cocoons; chromosome complement diploid with 16 metacentric chromosomes. Inter-specific molecular distances and their positions in the phylogenetic tree reveal that D. circumcisa and D. verrucula are clearly separated from their congeners.

Background Freshwater planarians of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) are distributed in a major part of the Old World and Australia, although until recently only very few species were known from China. Results Two new species of Dugesia from Southern China are described on the basis of an integrative taxonomic approach. BI and ML phylogenetic trees based on the independent genes and on the concatenated dataset had similar topologies, only differing in some nodes that were weakly supported. Phylogenetic trees based on the concatenated dataset revealed that D. adunca Chen & Sluys, sp. nov. and D. tumida Chen & Sluys, sp. nov. are not closely related and belong to different clades. The two new species occupy separate long branches with high support values and, thus, are well-differentiated from their congeners. Separate species status of D. adunca and D. tumida is supported also by the genetic distances between the species included in our analysis, albeit that COI distances varied greatly among species. Dugesia adunca from Guangxi Province is characterized by the following features: living mature animals rather small; asymmetrical openings of the oviducts into the bursal canal; penis papilla with shape of an aquiline bill, albeit with a blunt tip; asymmetrical penis papilla, with a large antero-dorsal lip and a much smaller ventro-posterior lip; very large seminal vesicle, provided with trabeculae; small diaphragm; mixoploid karyotype with diploid complements of 2n = 2x = 16 and triploid complements of 2n = 3x = 24, with all chromosomes being metacentric. Dugesia tumida from Guangdong Province is characterized by a penis papilla provided with a large, symmetrical penial valve from the middle of which arises the small, distal section of the papilla; a duct intercalated between the seminal vesicle and the small diaphragm; ventrally displaced ejaculatory duct curving upwards before opening to the exterior; penis papilla highly asymmetrical, having a slim and long ventral portion and a short and stubby dorsal part; vasa deferentia separately opening into antero-dorsal portion of seminal vesicle; oviducts openings symmetrically into ventral portion of the bursal canal, near its opening into the atrium; mixoploid karyotype, with diploid chromosome portraits of 2n = 2x = 16, and triploid complements of 2n = 3x = 24, with all chromosomes being metacentric. In the context of the various kinds of mixoploidy and the sexualization of specimens, reproductive modalities within the genus Dugesia are shortly discussed. Conclusion Molecular, morphological, and karyological markers show that the two populations examined represent members of the genus Dugesia and constitute two new, distinct species.