Reproductive modes and life cycles of freshwater planarians (Platyhelminthes, Tricladida, Paludicula) from southern Brazil

Abstract. Life cycles, ploidy levels, reproductive modes, and regeneration capacities of laboratory populations of the southern Brazil freshwater planarians Girardia tigrina and Girardia schubarti were studied. The mating behavior and life cycle of different populations from both species were analyzed regarding their karyotype, body size, modes of reproduction, and regeneration potential. Reproduction was evaluated according to fecundity, fertility, and fissiparity indices. In both species we observed that diploid planarians are ∼25% larger than triploid or mixoploid ones and that sexually reproducing populations have more offspring than asexual ones. Cocoon incubation time was shorter for G. tigrina , and G. schubarti showed a higher frequency of spontaneous malformations. G. tigrina was both more fecund (produced more cocoons) and more fertile (produced more hatchlings) than G. schubarti . The effects of alternative food sources (liver or egg yolk), demographic density, and water–salt concentration were evaluated in different populations of both species. All evaluated environmental factors had effects on fecundity and fertility indices and were more prominent for G. schubarti than for G. tigrina . Our data suggest that the populations of G. schubarti were more sensitive to environmental factors, and thus may be useful as bioindicators.     

Chromosome evolution in fish: BrdU replication patterns demonstrate chromosome homeologies in two species of the genus Astyanax

A comparison of R-banding patterns obtained by 5-bromodeoxyuridine incorporation was made between the chromosomes of two fish species of the genus Astyanax (Characiformes: Tetragonopterinae), A. altiparanae with 2n = 50 chromosomes, and A. schubarti with 2n = 36 chromosomes. The two species present the highest and the lowest chromosome numbers found in this fish genus, respectively, for which the modal chromosome number is 50. R-band homeology was detected, involving eleven chromosomes of A. schubarti and seventeen chromosomes of A. altiparanae, indicating a close chromosomal relationship between the two species, in spite of their great difference in chromosome number. A chromosome fusion in the past history of the group was hypothesized as a possible cause of the discrepant chromosome numbers of the two species.     

Repeated DNA elements in planarians of the Dugesia gonocephala group (Platyhelminthes,Tricladida)

The results of the isolation of repetitive DNA elements in the genome of Dugesia etrusca, a species of the Dugesia gonocephala group, are reported. These sequences, about 1.4 kb long, represent only a part of longer interspersed genomic units (De1 family) and appear to be limited to the genome of some planarians of this group, as indicated by a Southern blot analysis performed in different species and populations. The genomic relationships among different species and populations of the D. gonocephala group are discussed in relation to the results obtained in the present work. This revised version was published online in July 2006 with corrections to the Cover Date.     

Biogeography and karyotypes of freshwater planarians (Platyhelminthes, Tricladida, Paludicola) in southern Brazil

In the Tricladida (Platyhelminthes), the incidence of different biotypes identified by several ploidy levels is very common. Planarians collected in the State of Rio Grande do Sul were identified using cytogenetics. Different species distributions were observed with respect to Rio Grande do Sul's geomorphology, which could have been caused by their different microhabitats. Girardia tigrina and G. anderlani consisted of diploid and triploid individuals, whereas G. schubarti showed diploids, triploids, and mixoploids; for all these species, individuals of different ploidies were sympatric. Only for diploid G. anderlani were B chromosomes observed. These B chromosomes seem to have an irregular segregational behavior during mitosis, and possibly also during meiosis. However the processes (e.g., selection, mutation) of maintaining 2n, 3n, and 2n/3n individuals within natural populations of G. schubarti remain to be clarified.     

Resistance of two planarian species to UV-irradiation

The aim of this work was to determine the effects of 20, 25 and 30 minute UV-irradiation periods lambda = 253.5 nm to two planarian species Dugesia tigrina (Gir.) and Polycelis felina (Daly.). In vivo, UV light effects have been reported to affect intracellular receptors and disrupt simple behaviour. The effects of UV-rays on mortality and behavior as well as morphological, cytological and histological changes in the two planarian species were assessed, and the course and the dynamics of regenerative processes were compared between them. Experimental populations of Dugesia tigrina and Polycelis felina species were maintained in laboratory conditions at room temperature. Mortality, behavioral and morphological changes were monitored daily by means of a light stereomicroscope. For cytological and histopathological analysis, planarians were fixed in Bouine fixative on the first, second, third, fifth and seventh day after exposure to UV-irradiation, respectively. They were embedded in paraffin, cut on a microtome, stained with toluidin blue and embedded in Canada-balsam. UV-rays caused mortality, behavioral, morphological, cytological and histological changes in each planarian species. In regeneration of damaged body parts reticular cells and neoblasts played the main role. Neoblasts as totipotent cells extremely increased in number in the area of damaged tissue, immediately after UV-exposure. Dugesia tigrina was more sensitive to UV-rays than Polycelis felina due to possession of less pigmented cells. The course of regeneration in both species was similar. Most individuals of both species regenerated in 5 to 12 days after UV-irradiation.     

Regenerative and reproductive capacities of the fissiparous planarian Dugesia tahitiensis

A pilot study was performed to assess the regenerative capacities of Dugesia tahitiensis Gourbault, 1977, an exclusively fissiparous planarian species. Animals measuring 9.5–12.5 mm in length were used. Head regeneration rate determined by the appearance of eye spots (Brødsted, 1969: 29–46) was extremely high: at 23 °C, it took 43–59 h to regenerate clearly discernible eye spots in 28 specimens. For comparison, 3.8 days were reported for the regeneration of eye spots in 11.9–12.7 mm long D. tigrina (Girard) at 24 °C (Mead, 1985). As all posterior fragments regenerated a head, irrespective of the cutting level, D. tahitiensis seems to match the Phagocata velata (Stinger) type with a head frequency of 100% at every level (Teshirogi et al., 1977; cf. also Brøndsted, 1969:30).     

Karyology and biogeography of Dugesia japonica and Dugesia ryukyuensis in Kyûshû, southern Japan

In southern Japan, two dugesiid species of freshwater planarians are known: Dugesia japonica Ichikawa et Kawakatsu, 1964, and Dugesia ryukyuensis Kawakatsu, 1976. D. japonica is a common and polymorphic species widely distributed in the Far East (karyotypes: n=8, 2x=16, 3x=24). D. ryukyuensis is a species recorded only from the Southwest Islands of Japan (Nansei Shotô) (karyotypes: n=7, 2x=14, 3x=21). Recently, small populations of D. ryukyuensis were found in the lowland areas in Kyûshû on the East China Sea (the Nishisonogi Peninsula, the Shimabara Peninsula, the Gotô Islands, the Satsuma Peninsula, and the Ôsumi Peninsula). The current geographical distribution of D. japonica and D. ryukyuensis in southern Japan can be explained from geological and faunal viewpoints, as follows: (1) two separate inversions by the ancestor of D. japonica, one in the Miocene and one after early Quaternary; (2) only one expansion of its domain by the ancestor of D. ryukyuensis in the Miocene.     

The freshwater planarian Dugesia sicula Lepori from Sardinia (Platyhelminthes,Tricladida)

Fissiparous strains of freshwater triclads of the Dugesia gonocephala group were collected from 12 localities in Sardinia all situated not more than 5 kilometers from the coast. Some environmental factors and the sexual status of the specimens were noted at the time of collection. During the laboratory rearing 30% of individuals of each strain became sexual (ex-fissiparous individuals). All the examined strains showed common karyological and morphological characteristics suggestive of the species Dugesia sicula Lepori. The chromosome complement, which was a constant 27+2–3 B chromosomes, was classified as aneutriploid, due to its clearly documented characteristics. The fissiparous populations of D. sicula appear to have a high degree of tolerance to variations in environmental factors, especially temperature. Within the D. gonocephala group this species has the broadest distribution in the Mediterranean region.     

The genus Dugesia in Australia, with its phylogenetic analysis and historical biogeography (Platyhelminthes, Tricladida, Dugesiidae)

The new species Dugesia notogaea Sluys & Kawakatsu, sp. n. is described from Queensland, Australia, representing the first record of the genus Dugesia for that continent. A phylogenetic analysis of the 68 species of Dugesia resulted in the discovery of a new apomorphy for the genus, and a cladogram showing two major phyletic groups. The distributional range of one group has a western accent, whereas that of the other group has an eastern emphasis; the ranges of these two groups overlap in Africa and the Mediterranean region. We attempt to determine the ancestral area for both the Dugesiidae and Dugesia. A scenario for the historical biogeography of the genus Dugesia is discussed. In this scenario it is presumed that the geological history of the Australasian region enabled Dugesia's from southeast Asia to colonize northern Australia. © 1998 The Norwegian Academy of Science and Letters     

The preference for movement direction in a T-maze in planarians

Characteristics of spatial orientation in T-maze were studied in 1768 Planaria of following types: Dugesia tigrina (sexless and sexual race), Dugesia lugubris, Ijmia tenuis, Bdellacephala punctata. It was shown that from one third to one half of individuals were characterized by asymmetry of movement direction preference. The preference of right turning was typical for Dugesia tigrina; Dugesia lugubris, Ijmia tenuis, Bdellacephala punctata preferred left turning. The asymmetry described is considered as a primitive form of species functional asymmetry.     

中国淡水三角涡虫(Dugesia sp)的染色体研究(Ⅰ)

利用空气干燥法对不同产地淡水三角涡虫的染色体进行了研究。核型分析表明：河南淇县鱼泉三角涡虫(Dugesia sp)和浙江杭州龙井三角涡虫(Dugesia sp)体细胞中有16条染色体,为二倍体,核型公式为2n=2x=16=16m,均为具中部着丝粒染色体;河南济源不老泉三角涡虫(Dugesia sp)有24条染色体,为三倍体,核型公式为2n=3x=24=24m,亦全部由中部着丝粒染色体组成。上述3个产地淡水三角涡虫染色体的形态较为接近。北京樱桃沟三角涡虫(Dugesia sp)的体细胞染色体数目为24,为三倍体,核型公式为2n=3x=24=22m 2sm,由中部和亚中部着丝粒染色体组成,其中第2、4号各有一条染色体属于亚中部着丝粒染色体。研究结果表明：4个产地三角涡虫的体细胞染色体数目存在较大差异,包括二倍体(2n=2x=16)和三倍体(2n=3x=24),染色体基数属于x=8类型。     

Identification of small non-coding RNAs in the planarian Dugesia japonica via deep sequencing

Freshwater planarian flatworm possesses an extraordinary ability to regenerate lost body parts after amputation; it is perfect organism model in regeneration and stem cell biology. Recently, small RNAs have been an increasing concern and studied in many aspects, including regeneration and stem cell biology, among others. In the current study, the large-scale cloning and sequencing of sRNAs from the intact and regenerative planarian Dugesia japonica are reported. Sequence analysis shows that sRNAs between 18nt and 40nt are mainly microRNAs and piRNAs. In addition, 209 conserved miRNAs and 12 novel miRNAs are identified. Especially, a better screening target method, negative-correlation relationship of miRNAs and mRNA, is adopted to improve target prediction accuracy. Similar to miRNAs, a diverse population of piRNAs and changes in the two samples are also listed. The present study is the first to report on the important role of sRNAs during planarian Dugesia japonica regeneration.     

Chromosome bands in freshwater triclads

Four species of Triclads belonging to three families, Dugesia polychroa and Dugesia mediterranea (Dugesiidae), Planaria torva (Planariidae) and Dendrocoelum lacteum (Dendrocoelidae) were studied for chromosome banding: C-banding for all the species, ASG banding for Dugesia polychroa and G-banding for Dugesia polychroa and Dugesia mediterranea. C-banding results for Dugesia lugubris-polychroa group suggest a high level of heterochromatin evolution in the group. Small bands strictly limited to the centromeric region were seen in Dugusia lugubris and this pattern is similar to the patterns of some species belonging to other families, notably Planaria torva and Dendrocoelum lacteum. This could be considered a primitive character. In contrast numerous heterochromatin bands which are useful to characterize the different karyotypes were seen in Dugesia polychroa and Dugesia mediterranea. Our data suggest that heterochromatin is important in freshwater triclad speciation.     

On the taxonomic status of a Dugesia species from Buenos Aires (Platyhelminthes: Turbellaria: Tricladida)

The taxonomic and distributional statuses of the freshwater planarians of South America are imperfectly understood, and a comprehensive review is sorely needed. The species known from the vicinity of Buenos Aires, Argentina, was first described by Böhmig (1902) under the name Planaria dubia Borelli, 1895. Subsequent authors (Kenk, 1930, 1974; Cazzaniga & Curino, 1987) have cast doubt on this assignation. Some have assigned it to Dugesia anceps Kenk; Hyman (1959) was inclined to regard the Buenos Aires planarian as a geographical variant of Dugesia sanchezi Hyman. A comparison of D. sanchezi from the mountainous region of central Chile with specimens from Buenos Aires confirms that the Argentinian material is a distinct species.The differences between the two species are subtle. The Argentinian specimens are distinctive in their possession of infranucleate epithelia of the atria, penis, and bursal canal, as well as in their penial structure. In both species, the epithelia of the atria and bursal canal are glandular. Those of D. sanchezi are nucleate and their secretions are cyanophilous. In the Argentinian specimens, these same epithelia are infranucleate; but while the secretion of the bursal canal is indeed cyanophilous, that of the atrium is erythrophilous. The presence, in these same specimens, of cyanophilous glands opening into the penial vesicles, and of erythrophilous glands whose secretions contribute to a viscous plug in the ejaculatory duct, suggests a difference in the physiology of copulation.It is apparent that the Argentinian specimens are distinct from D. sanchezi Hyman, and probably they represent a new species, one to be described in detail elsewhere. In any case, Hyman's (1959) suggestion that D. sanchezi occurs near Buenos Aires is mistaken; so also is Cazzaniga's & Curino's (1987) identification of specimens from Buenos Aires as D. anceps.     

Feeding on gastropods by lake-dwelling Polycelis in the absence and presence of Dugesia polychroa (Turbellaria, Tricladida)

SUMMARY. Using a serological technique, the extent of feeding upon snails by Polycelis tenuis was compared in habitats lacking and containing Dugesia polychroa which is the major triclad predator of snails. It was found that P. tenuis ate significantly more snails when D. polychroa was absent. It was concluded that in the field interspecific competition for snails between these two triclad species was a more powerful process than any social feeding by P. tenuis on snails captured by D. polychroa .
Feeding on snails as a group by P. tenuis and P. nigra in the habitat where the two triclads co-existed was of a similar magnitude and this was also true of their feeding on the common snail species and genera. Approximately 10% of the triclads gave a positive reaction to snail antisera.
Feeding on snails by P. nigra in the three habitats lacking D. polychroa was similar in extent and did not differ from that of P. tenuis . It was concluded that P. nigra would be similarly affected by the presence and absence of D. polychroa as shown for P. tenuis .
No definitive seasonal pattern was detected in the incidence of feeding upon snails by either Polycelis species but this may have been obscured by seasonal changes in the detection period of snail antigens due to temperature effects. There was some evidence that such feeding increased during the period when small snails were present in the population.     

A comparative study of the regenerative processes in a trematode, Philophthalmus gralli, and a planarian, Dugesia dorotocephala

Adults of Philophthalmus gralli, an eyefluke of birds, were laterally amputated mid-way between the anterior testis and the ventral sucker. Worms were sampled at various short time intervals (30 min-6 h) after in vitro culture and long time intervals (1-8 days) after transplantation back to the host. Specimens were fixed for scanning electron microscopy and light microscopy and compared to the planarian, Dugesia dorotocephala, which was laterally transected in the pharyngeal region and maintained in springwater. It was found that wound closure took place by 2 days in the planaria; however, at the end of 8 days wound closure had been initiated but not yet completed in P. gralli. Replacement of major tissues was observed only in D. dorotocephala. Because calcium had been reported to be critical for planarian regeneration, a histochemical stain for calcium distribution was carried out. At the end of the 2-day study, no differences could be found in calcium distribution between the two organisms. No areas of calcium concentration were noted in any tissue important for the regenerative process.