Integrative description of a new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Shennongjia, Central China.

A new species of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Xiangxi River, Shennongjia Forestry District, Hubei Province, China, is described on the basis of an integrative approach, involving morphology, and molecular systematics. The new species Dugesia saccaria A-T. Wang & Sluys, sp. nov. is characterized by the following features: a dumb-bell-shaped, muscularized hump located just anterior to the knee-shaped bend in the bursal canal; a ventrally displaced ejaculatory duct, which, however, opens terminally through the dorsal portion of the blunt tip of the penis papilla; a ventrally located seminal vesicle, giving rise to a vertically running duct that eventually curves downwards to communicate with the ejaculatory duct via a small diaphragm; oviducts opening asymmetrically into the dorsal portion of the common atrium and at the knee-shaped part of the bursal canal. The phylogenetic position of the new species was determined using four molecular markers (18S rDNA; ITS-1; 28S rDNA; COI), which suggested that it groups with other species of Dugesia from the Australasian and Oriental biogeographical regions.

Molecular phylogenetics facilitates the first historical biogeographic analysis of the hammerhead worms (Platyhelminthes: Tricladida: Bipaliinae), with the description of twelve new species and two new genera.

The hammerhead worms constitute a land planarian subfamily (Platyhelminthes: Tricladida: Bipaliinae) that is popular among natural historians as well as citizen scientists due to their characteristic semi-lunar-shaped head, the striking colours of many species, and the worldwide presence of introduced species, which has raised much concern about their putative impact on the receiving ecosystems. Despite such current awareness, the majority of Bipaliinae species was described before the second half of the 20th century. Over the past few decades, description of new species has been rather scarce. In the present paper, we describe twelve new species and two new genera from Madagascar, Borneo, and Japan, mainly using specimens that form part of the collections of Naturalis Biodiversity Center, Leiden. Species identification has been based on morphology, inner anatomy, and, when available, molecular information. Our molecular phylogenetic tree suggests that either Malagasy or Bornean representatives form the sister-group to the rest of Bipaliinae. The phylogeny suggests also that the Bipaliinae comprises several new and, as yet, undescribed genera. Although the geographical origin of the subfamily is uncertain and may involve either Madagascar or the Bornean region, a molecular time-calibration of the phylogenetic tree indicated that the origin of the Bipaliinae may date back to about 253 Mya, placing its origin near the Permian-Triassic transition and, thus, on Pangea.

Two new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from the subtropical monsoon region in Southern China, with a discussion on reproductive modalities.

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.

Integrative taxonomy unveils a new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from the southern portion of the Taihang Mountains in northern China, with the description of its complete mitogenome and an exploratory analysis of mitochondrial gene order as a taxonomic character.

A new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from northern China is described on the basis of an integrative approach, involving morphology, karyology, histology, molecular distance, molecular phylogeny, and mitochondrial gene order. Here, we present the complete mitogenome of the new species Dugesia constrictiva Chen & Dong, sp. nov. This new species is mainly characterized by the presence of the following features: asymmetrical openings of the oviducts; large, cuboidal copulatory bursa; vasa deferentia opening through the ventro-lateral wall of the seminal vesicle; laterally compressed seminal vesicle; ventrally displaced ejaculatory duct, opening at the blunt tip of the penis papilla; long duct intercalated between seminal vesicle and diaphragm; chromosome complement diploid, with 16 metacentric chromosomes; mitochondrial gene order as follows: cox1-E-nad6-nad5-S2-D-R-cox3-I-Q-K-atp6-V-nad1-W-cox2-P-nad3-A-nad2-M-H-F-rrnS-L1-Y-G-S1-rrnL-L2-T-atp8-C-N-cob-nad4l-nad4. In triclads, mitochondrial gene order is considerably conserved between freshwater planarians and land flatworms, whereas it is variable between marine planarians and both freshwater and land flatworms. The secondary structures of tRNAs are all equipped with 4 arms, excepting tRNA S1 and tRNA S2, which lack the D arm and have excessively enlarged loops. Numerous transpositions of tRNA are present between D. constrictiva and its congeners. Mitochondrial gene arrangements may form a new, additional tool for taxonomic studies. The phylogenetic tree based on analysis of the mitochondrial genome basically corroborates current classification of the higher taxa of planarian flatworms.

Two new species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from the tropical monsoon forest in southern China.

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 Dugesiacircumcisa 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, Dugesiaverrucula 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.

Integrative descriptions of two new species of Dugesia from Hainan Island, China (Platyhelminthes, Tricladida, Dugesiidae).

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.

First records of two alien land flatworms (Tricladida, Geoplanidae) from Northeastern Italy.

During a period of intense rainfall (May 2019), several specimens of land flatworms were collected from a private garden in Palazzolo dello Stella (Friuli Venezia Giulia, Udine, Italy: 45°47'40.5"N, 13°05'17.2"E). Planarians were found both in a cultivated part of the garden and in a part covered with gravel and with trees and shrubs (Pyracantha sp., Olea europaea, Pyrus communis). The animals were observed under branches, stones, tufa blocks, and pots close to a small artificial pond, but also in other parts of the garden, as well as inside buildings.

Cryptic species delineation in freshwater planarians of the genus Dugesia (Platyhelminthes, Tricladida): Extreme intraindividual genetic diversity, morphological stasis, and karyological variability.

The keystone of planarian taxonomy traditionally has been the anatomy of the copulatory apparatus. However, many planarian species comprise asexual fissiparous populations, with the fissiparous animals not developing a copulatory apparatus, thus precluding their morphological identification. Incorporation of molecular data into planarian systematics has been of great value, not only in the identification of fissiparous individuals but also as an additional source of information for determining species boundaries. Nevertheless, the discrepancy between morphological and molecular data has highlighted the need for extra sources of taxonomic information. Moreover, a recent study has pointed out that fissiparous reproduction may lead to high levels of intraindividual genetic diversity in planarians, which may mislead molecular analyses. In the present study we aim to test a new up-to-date integrative taxonomic procedure for planarians, including intraindividual genetic data and additional sources of taxonomic information, besides morphology and DNA, using Dugesia subtentaculata sensu lato as a model organism, a species with an intricate taxonomic history. First, we used three different methods for molecular species delimitation on single locus datasets, both with and without intraindividual information, for formulating Primary Species Hypotheses (PSHs). Subsequently, Secondary Species Hypotheses (SSHs) were formulated on the basis of three types of information: (1) a coalescent-based species delimitation method applied to multilocus data, (2) morphology of the copulatory apparatus, and (3) karyological metrics. This resulted in the delimitation of four morphologically cryptic species within the nominal species D. subtentaculata. Our results provide evidence that the analysis of intraindividual genetic data is essential for properly developing PSHs in planarians. Our study reveals also that karyological differentiation, rather than morphological differentiation, may play an important role in speciation processes in planarians, thus suggesting that the currently known diversity of the group could be highly underestimated.

Phylogeny and biogeography of the Cavernicola (Platyhelminthes: Tricladida): Relicts of an epigean group sheltering in caves?

The planarian suborder Cavernicola Sluys, 1990 was originally created to house five species of triclad flatworms with special morphological features and a surprisingly discontinuous and broad geographic distribution. These five species could not be accommodated with any degree of certainty in any of the three taxonomic groups existing at that moment, viz., Paludicola Hallez, 1892, Terricola Hallez, 1892, and Maricola Hallez, 1892. The scarce representation of the group and the peculiarities of the morphological features of the species, including several described more recently, have complicated new tests of the monophyly of the Cavernicola, the assessment of its taxonomic status, as well as the resolution of its internal relationships. Here we present the first molecular study including all genera currently known for the group, excepting one. We analysed newly generated 18S and 28S rDNA data for these species, together with a broad representation of other triclad flatworms. The resulting phylogenetic trees supported the monophyly of the Cavernicola, as well as its sister-group relationship to the Maricola. The sister-group relationship to the Maricola and affinities within the Cavernicola falsify the morphology-based phylogeny of the latter that was proposed previously. The relatively high diversity of some cavernicolan genera suggests that the presumed rarity of the group actually may in part be due to a collecting artefact. Ancestral state reconstruction analyses suggest that the ancestral habitat of the group concerned epigean freshwater conditions. Our results point to an evolutionary scenario in which the Cavernicola (a) originated in a freshwater habitat, (b) as the sister clade of the marine triclads, and (c) subsequently radiated and colonized both epigean and hypogean environments. Competition with other planarians, notably members of the Continenticola, or changes in epigean habitat conditions are two possible explanations -still to be tested- for the loss of most epigean diversity of the Cavernicola, which is currently reflected in their highly disjunct distributions.

Two new species of marine flatworm from southern China facilitate determination of the phylogenetic position of the genus Nerpa Marcus, 1948 and the histochemical structure of the nervous system in the genus Paucumara Sluys, 1989 (Platyhelminthes, Tricladida, Maricola).

Two new species of flatworm, collected from a beach at eastern Shenzhen, China, were studied through an integrative approach by combining morphological, histological, histochemical (acetylcholinesterase, AChE), and molecular (18S r- DNA) data. These species belong to two genera of marine triclads, previously unrecorded from China, viz. Nerpa Marcus, 1948 and Paucumara Sluys, 1989.        Nerpa fistulata Wang Chen, sp. nov. is characterized by: transparent body; principally pentamerous intestine with three distinct commissures; two very large, prepharyngeal testis follicles; a semi-circular lens in each eye cup; a penis papilla provided with a chitinized, pointed stylet; lateral bursae communicating with the oviduct and opening ventrally to the exterior via a duct. Phylogenetically N. fistulata groups with one member of the family Bdellouridae. This new, Chinese species of Nerpa introduces a major geographic disjunction, as the type species N. evelinae was described from the bay of Santos, Brazil, so that the genus is now known from both Atlantic as well as Pacific coasts.        The species Paucumara falcata Wang Li, sp. nov. is characterized by: three distinct pale yellow transverse pigmentation bands on its dorsal side, between which some snowflake-like specks are randomly distributed, and a brown transverse band anteriorly to the eyes; 8-11 testicular follicles on either side of the body, the follicles extending from immediately behind the ovaries to half-way along the pharyngeal pocket; a musculo-parenchymatic organ with a sclerotic, curved tip projecting from the anterior wall of the male atrium, ventrally to the root of the penis papilla. Phylogenetically P. falcata groups with its congener P. trigonocephala, with the genus Paucumara forming the sister taxon of the genus Ectoplana. Comparison of the nerve structure of P. falcata, as revealed by AChE histochemistry, with that of eight other species of triclad suggested that the nervous system of marine planarians is simpler than that of species of freshwater planarians, but revealed also that the nerve structure is rather variable among species.        The copulatory position exhibited by two partners in Paucumara falcata is remarkable in that they intertwine, with their heads pointing downwards and the tails pointing upwards, the entire process lasting about 10 min. Such a copulatory position has never before been reported for triclad flatworms.

A new genus and species of a terrestrial cavernicolan planarian from Barro Colorado Island, Panama (Platyhelminthes, Tricladida, Cavernicola).

The paper provides a taxonomic description of a new genus and species of terrestrial cavernicolan triclad flatworm from Barro Colorado Island, Panama. Molecular sequences of this species were already deposited in GenBank several years ago under the provisional taxon name Cavernicola sp. Animals have the following characteristics: absence of pigmentation and eyes; mouth opening located halfway in the pharyngeal cavity; few, ventral testes extending from a position far posterior to the ovaries to directly behind the pharyngeal cavity; strongly recurving sperm ducts, thus forming a loop; horizontally oriented cone-shaped penis papilla; common oviduct oriented perpendicular to the bursal canal and opening into the latter shortly before the canal communicates with the copulatory bursa; sac-shaped copulatory bursa lacking a distinct, single lumen and filled with a mass of syncytial cells, with interspersed nuclei.

New molecular sequences for two genera of marine planarians facilitate determination of their position in the phylogenetic tree, with new records for two species (Platyhelminthes, Tricladida, Maricola).

For the first time, molecular sequences of the 18S ribosomal DNA were generated for representatives of the genera Obrimoposthia Sluys & Ball, 1989 and Paucumara Sluys, 1989 of the suborder of the marine triclads, or Maricola, by analyzing the species Obrimoposthiawandeli (Hallez, 1906) and Paucumaratrigonocephala (Ijima & Kaburaki, 1916). On the basis of this molecular data the phylogenetic position of these two genera in the phylogenetic tree of the Maricola was determined and compared with their position in the phylogeny based on the analysis of anatomical features. New records for these two species are documented and their taxonomic status is determined on the basis of histological studies.

Planarian Diversity and Phylogeny.

Hundreds of planarian species exist worldwide, representing a rich phenotypic diversity. This chapter presents an overview of the morphology and anatomy of various taxonomic groups of planarian flatworms, focusing on features enabling recognition and identification of the animals. The most recent view on the phylogenetic relationships of the planarians is presented, together with geographic distribution patterns of major groups of triclads. The chapter concludes with a brief methodological section outlining species identification on basis of anatomical features. In conjunction with the established laboratory model species, the phenotypic diversity of planarians provides rich opportunities for comparative studies, which this chapter aims to inspire.

Basic Histological Techniques for Planarians.

The chapter provides a concise account of collecting techniques, and basic histological techniques for investigating planarians, intended for use by practicing scientists and postgraduate students new to the field. Selected hands-on methods are described that will lead to well-stained preparations suitable for light microscopic examination for subsequent taxonomic identification or general investigations into planarian functional cytology and histology. Major equipment items that are minimal essential requirements of a functional histological laboratory are indicated. The chapter includes sectioning and staining troubleshooting guides that will be very helpful for those who must work in the laboratory without the opportunity to consult qualified and experienced technicians or microscopists.

Land flatworms (Platyhelminthes, Geoplanidae) of São Tomé: a first account on their diversity, with the description of five new species.

The present contribution provides the first faunistic and taxonomic account of six species of land flatworm from the island of São Tomé, including five new species of the genus Othelosoma Gray, 1869 and the introduced Bipalium kewense Moseley, 1878. One of the new species represents the first African land flatworm that has specks on its dorsal body surface, instead of stripes or a more or less uniform colouration. At least two of the new species were observed to prey on snails. The study details the fourth record of a sclerotic spermatophore in a species of land flatworm, and discusses the definition and homology of double female genital canals in African and Indian species of the genus Othelosoma.

Bipolarity in planarians is not induced by space travel.

Available evidence strongly suggests that alternative, Earth-bound explanations should be sought first for the occurrence of a single bipolar planarian flatworm returning from space travel. Double-headed worms have been amply documented as arising under experimental conditions as well as spontaneously in stock cultures of planarians.

First report of the land planarian Diversibipalium multilineatum (Makino & Shirasawa, 1983) (Platyhelminthes, Tricladida, Continenticola) in Europe.

Introduction of alien species may significantly affect soil ecosystems, through predation or disruption of components of native ecosystems (Winsor et al. 2004; Álvarez-Presas et al. 2014; Justine et al. 2014). Land planarians have been reported as alien species in soils throughout the world and, among those, some species are considered to be successful invaders, e.g. Platydemus manokwari de Beauchamp, 1963, Arthurdendyus triangulatus (Dendy, 1894), Bipalium adventitium Hyman, 1943, Bipalium kewense Moseley, 1878 and Dolichoplana striata Moseley, 1877 (Winsor et al. 2004; Álvarez-Presas et al. 2014; Justine et al. 2014, 2015). Soil moisture status seems to be an important element for their successful invasion (Fraser & Boag 1998). In Europe at least 18 species of alien land planarians have been recorded since now and some of them are considered as invasive ones, e.g. P. manokwari (cf. Justine et al. 2014). Although the alien land planarian B. kewense has been reported to occur in many greenhouses in Italy (Bello et al. 1995), no data are available on its establishment and/or impact on natural environments. On 28th September 2014, 20 specimens (~1 individual/m2) of the land planarian Diversibipalium multilineatum (Makino & Shirasawa, 1983) (Fig. 1), native to Japan, were collected under pots, branches and plastic materials in a private garden located in the center of Bologna (Emilia Romagna, Central Italy), near the urban park Giardini Margherita (44°29' N, 11°21' E; WGS84). Thirty plant species (both indigenous and alien), mainly cultivated as bonsai (e.g. Lagerstroemia indica L., Juniperus procumbens (Siebold ex Endl.) Miquel), were present in this shady, wet garden (25 m2). Between March 2014 and June 2015, 70 more specimens of D. multilineatum were collected at the same site, mainly at dusk and dawn after rain. Reproduction by fission and regeneration processes were observed in several of those specimens, which were kept for some time in captivity. A specimen of D. multilineatum was also collected in a garden in Léguevin (Haute-Garonne, France), which will be described in a forthcoming paper by Justine et al. (in prep.) (see also Kawakatsu et al. 2014). Specimens without a genital pore were initially ascribed to D. multilineatum on the basis of their external appearance: the dorsal surface was brownish yellow and presented five longitudinal stripes at the head plate and the neck, showing the typical appearance of the species. The middorsal stripe was widened at its anterior end, on the head plate, and at the pharynx level. The ventral pattern of the animals at the pharyngeal region was also characteristic, with the middorsal stripe widened at this level. The Italian Diversibipalium specimens used for the molecular analysis were fixed and preserved in absolute ethanol. Fragments of the mitochondrial gene COI and 28S ribosomal RNA nuclear gene (GenBank Acc. Numbers KU245358 and KU245357, respectively) were obtained using the procedure and COI primers described in Álvarez-Presas et al. (2008) and Solà et al. (2013). The French specimen's COI (Specimen MNHN JL177, GenBank Acc. Number KT922162) was obtained as described in Justine et al. (2015). 28S sequences of 14 Bipaliinae specimens and four Microplana species (outgroup) retrieved from GenBank were included in the phylogenetic analyses (Fig. 2). Sequence alignment was obtained by using the online software MAFFT version 7 (Katoh & Standley 2013), while ambiguously aligned positions were removed using the program Gblocks (Talavera & Castresana 2007) with default settings, excepting the minimum number of sequences for a flank position at the minimum value (set at 10) and with half of the allowed gap positions. The final alignment had a length of 1589 bp. We used two phylogenetic inference approaches: maximum likelihood (ML), using the RaxML 8.2.3 software (Stamatakis 2014), and Bayesian inferences (BI), using MrBayes 3.2.4 (Ronquist et al. 2012). The evolutionary model used, GTR+I+G, was estimated to be the best with the software jModeltest 2.1.7 (Darriba et al. 2012; Guindon & Gascuel 2003), using the Akaike Information Criterion (AIC). MrBayes analyses were performed for 10-milion generation with sampling parameters every 103 and a 25% default burn-in value for the final trees. Convergence of the two runs (average standard deviation of split frequencies << 0.01) and likelihood stationarity were checked. The maximum likelihood analyses were performed under 1000 bootstrap pseudoreplicates. The phylogenetic results show a close and highly supported relationship of the Italian Diversibipalium specimens with those from Japan and South Korea that have been identified as D. multilineatum (Fig. 2). Diversibipalium multilineatum is the sister-group of B. nobile Kawakatsu & Makino, 1982, but with low support. The COI sequences of the French (MNHN JL177) and the Italian Diversibipalium specimens were compared in Geneious v. 8.0.5 (http://www.geneious.com, Kearse et al. 2012) and were found to be identical. These results indicate that the species introduced in both countries is the same, and most probably concerns the species D. multilineatum. The pathways of introduction of D. multilineatum are currently unknown, although a relationship between the horticultural trade and the introduction of alien land planarians is well known (Álvarez-Presas et al. 2014 and references therein). Here we report the first occurrence of individuals of D. multilineatum outside Asia. The GenBank sequence of D. multilineatum from South Korea is not yet supported by a published description of the specimen, while it is debatable whether South Korea should be considered part of the natural range of D. multilineatum, which only seems to include Japan. In the present paper, we consider the South Korean animal to be an introduced specimen. Soil moisture status, temperature, and food availability are considered to be the main factors determining the presence of terrestrial planarians (Boag et al. 1998); the microclimatic conditions of the Italian garden were similar to plant nurseries and greenhouses, while an abundance of food was available, such as isopods [Porcellionides pruinosus (Brandt, 1833)], oligochaetes [Dendrobaena attemsi (Michaelsen, 1902) and several juveniles of Lumbricus spp.] and gastropods [Cernuella cisalpina (Rossmassler, 1837), Cornu aspersum (O.F. Müller 1774), Deroceras reticulatum (O.F. Müller, 1774), Discus rotundatus (O.F. Müller, 1774), Limacus flavus (Linnaeus, 1758), Milax nigricans (Philippi, 1836), Papillifera papillaris (Linnaeus, 1758), Pomatias elegans (O.F. Müller, 1774)]. Moreover, winter 2014 reached the highest temperatures and rainfall of the last two decades (source: CNR-ISAC, Bologna), thus favouring establishment and spread of D. multilineatum. The potential environmental impacts of some invasive flatworms are well documented (Álvarez-Presas et al. 2014; Justine et al. 2014) and, even if these effects have not yet been assessed for D. multilineatum, the adoption of precautionary measures and of early intervention is here strongly recommended (Genovesi & Shine 2004). Finally, knowledge of the introduction pathway(s), together with the analysis of prey preference and possible impact on the invertebrate fauna, will be essential to halt or at least to limit the spread of this introduced land flatworm.

The taxonomic status of Dugesiabiblica from Israel and Turkey (Platyhelminthes, Tricladida, Dugesiidae).

The taxonomic status of Dugesiabiblica (Platyhelminthes, Tricladida, Dugesiidae) from Israel and Turkey is problematic due to its morphological similarity with Dugesiasicula since these nominal species present overlapping characters. In this study we analyzed histological preparations of specimens of these two nominal species and also compared mitochondrial COI gene sequences from Israeli populations to the already known haplotype composition of Dugesiasicula. We concluded that these animals belong to the same species and therefore we consider Dugesiabiblica to be a junior synonym of Dugesiasicula. This implies that the distribution range of Dugesiasicula is even wider than previously thought, and that the species is present all around the Mediterranean Basin and on many of its islands.

A new and aberrant species of Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Madagascar.

In this paper we report a new species of Dugesia of the family Dugesiidae from Madagascar, representing the fourth species of freshwater planarian known from this global biodiversity hotspot. In some respects the new species is aberrant, when compared with its congeners, being characterized by a head with smoothly rounded auricles, a peculiar course of the oviducts, including the presence of a common posterior extension, and by the asymmetrical openings of the vasa deferentia at about halfway along the seminal vesicle. Further, it is characterized by a ventral course of the ejaculatory duct with a terminal opening, very long spermiducal vesicles and unstalked cocoons. Its diploid chromosome complement with 18 chromosomes represents an uncommon feature among fissiparous species of Dugesia.

Integrative taxonomy of a new species of planarian from the Lake Ohrid basin, including an analysis of biogeographical patterns in freshwater triclads from the Ohrid region (Platyhelminthes, Tricladida, Dugesiidae).

A new species of the genus Dugesia is described from the Lake Ohrid region in the western part of the Balkan Peninsula, forming the first fully documented species description for this genus in the Ohrid area. The morphological species delimitation is supported by complementary molecular, karyological, and cytogenetic data available from the literature. Therefore, species delineation is based on a truly integrative approach. Further, a short account on the degree of freshwater planarian endemicity in the Ohrid region is provided.