Investigation of the ultrastructure of Dendrocoelum constrictum (Platyhelminthes,Tricladida) spermatogenesis and mature spermatozoa

To add to our understanding of dendrocoelid spermatozoa and to describe additional phylogenetic characters, the ultrastructure of the testis was investigated in the subterranean freshwater planarian Dendrocoelum constrictum. This is the first study investigating spermatogenesis and spermatozoon ultrastructure in a subterranean freshwater planarian species. We found that the basic structure of spermatozoa in D. constrictum is similar to that of other Tricladida that have been studied previously. In fact, D. constrictum spermatozoa possess an elongated nucleus, one giant mitochondrion, and two subterminal flagella with a 9 + ‘1’ pattern. The flagella emerge together from one side of the spermatozoon. However, D. constrictum has some characteristics that have not yet been described for other freshwater planarians. In fact, the number of cortical microtubules reaches the maximum number in the anterior and middle part of region I, and then decrease until they disappear towards the posterior extremity of the spermatozoon. The extreme tip of the anterior region of the spermatozoon exhibits a specific external ornamentation of the plasma membrane.     

Genetic differences among several species of Tricladida from the relict Lake Ohrid as revealed by enzyme electrophoresis

Six endemic and two widely distributed species living in Lake Ohrid were studied. In general, these hermaphroditic animals displayed no signs of departure from Hardy-Weinberg equilibrium. Genetic variation in all but one of the endemic species was of the same extent as that in geographically wide ranging invertebrates. On the other hand, the Lake Ohrid population of the common European species Dendrocoelum lacteum was monomorphic at all loci examined. D. sanctinaumi, one of the endemic species, exhibited a clear genetic subdivision into spring and littoral subpopulations. The genetic differentiation of Crenobia alpina alpina and C. a. montenigrina proved commensurable to that of well separated species from other genera. The data suggest that the separation of particular lineages in the set of Lake Ohrid endemics was widely dispersed over time.     

Patterns of genetic differentiation and population history of endemic isopods (Asellidae) from ancient Lake Ohrid: combining allozyme and mtDNA data

Ancient lakes as places of extensive speciation processes have been characterized by a high degree of endemicity and biodiversity. The most outstanding European ancient lake is the oligotrophic and karstic Balkan Lake Ohrid. The lake is inhabited by a number of endemic species, but their evolutionary history is largely unresolved. in the present study, the genetic structure, gene genealogy and demographic history of the representatives of the Ohridian endemic Proasellus species were studied using both biparentally (allozyme loci) and maternally (partial mitochondrial cytochrome oxidase subunit I gene) inherited markers. Both data sets gave similar results and supported discrepancies among genetic differentiation, the current morphology-based taxonomy and bathymetric segregation. Horizontal distribution of endemic Proasellus species (Lake Ohrid vs adjacent feeder springs) within the lake presumably promote parapatric speciation whereas the main role of vertical barriers into diversification processes was not fully supported. The analyses of demographic history suggested the decline of endemic isopod populations. The radiation of endemic Proasellus populations within the lake could have started from the sublittoral/profundal zone towards the littoral or in the opposite direction — from the littoral to the profundal. Our analyses did not exclude both possibilities.     

Rapid evolution of an ancient lake species flock: Freshwater limpets (Gastropoda: Ancylidae) in the Balkan Lake Ohrid

Ancient lakes have long been recognized as evolutionary theatres and hot spots of endemism; the evolution of their morphologically often highly diverse species flocks has received much attention. However, as each ancient lake has its own geological and evolutionary history, modes of speciation may differ from system to system. Ancient lakes can act as evolutionary reservoirs that assure the survival of relict species, but at the same time extant species may evolve through intralacustrine speciation. Other aspects of interest are the actual rates of immigration, diversification or extinction as well as the temporal framework of morphological change. Many of these questions have been addressed in the African (e.g. Lake Tanganyika) and Asian (e.g. Lake Baikal) ancient lakes. For an European ancient lakes (e.g. Lakes Ohrid and Prespa), such studies are largely missing. In the present paper, extraordinarily shaped endemic freshwater limpets of the genus Ancylus from the Balkan Lake Ohrid are used in a phylogeographic and phylogenetic context to test whether they represent an ancient lake species flock, to study the mode of speciation, and to assess the timing of morphological change. Based on DNA data from two mitochondrial genes (COI, LSU rDNA), it has been found that the Lake Ohrid Ancylus species form an endemic monophyletic group. In addition, the lake's feeder springs are inhabited by another, undescribed Ancylus species. All other studied waterbodies within the watershed do not support their own Ancylus lineages but are inhabited by a widespread Mediterranean taxon. The split between the species endemic to the lake and its sister taxon is dated to 1.4±0.6 million years ago. The study presents the first genetic confirmation for the existence of a species flock in a European ancient lake. Contrary to the prevailing opinion it shows that, concerning Ancylus, Lake Ohrid represents a site of intralacustrine speciation rather than an evolutionary reservoir. Moreover, it provides the first evidence for rapid morphological change in an European ancient lake species flock. See also Electronic Supplement at: http://www.senckenberg.de/odes/06-12.htm.     

Phylogenetic relationships in Blumea (Asteraceae: Inuleae) as evidenced by molecular and morphological data

The present study, based on sequences of cpDNA (trnL-F & psbA-trnH) and nrDNA (ITS) and morphology, examined the evolutionary relationships in Blumea and its position among related genera. The results confirmed that the closest relatives of Blumea are Caesulia, Duhaldea and Pentanema p.p., and showed that the monotypic genera Blumeopsis and Merrittia are nested within Blumea. In Blumea s.l., two major, well-supported clades were recognised and a single species, the widespread Blumea balsamifera, that could not be placed with certainty relative to the two main clades. The two main clades differ in habit, ecology and distribution. The Blumea densiflora clade contains shrubs and subshrubs of evergreen forests, distributed from continental Asia to New Guinea and Polynesia, whereas the Blumea lacera clade is a widespread paleotropical group that comprises mostly annual, weedy herbs of open forests and fields.     

Phylogenetic relationship and taxonomic position of Xylanthemum tianschanicum (Krasch.) Muradyan (Compositae,Anthemideae) as inferred from nrDNA ITS data

The central Asian endemic Xylanthemum tianschanicum (Krasch.) Muradyan (Compositae, Anthemideae) is the only species of Xylanthemum Tzvelev with radiate capitula. While the species was formerly found to be closely related to members of the subtribe Handeliinae, other representatives of the genus were considered to be similar to members of Tanacetum. In order to assess its phylogenetic relationships and taxonomic classification, a molecular analysis based on ITS sequences of 13 representatives of the subtribe Handeliinae and 17 species of three other subtribes (Anthemidinae, Artemisiinae and Matricariinae) was performed. Due to the close relationship of the X. tianschanicum to the type species of Richteria (Richteria pyrethroides) in a well supported clade (PP = 1, BS = 100%), which is supported by morphological features concerning the indumentum, capitulum morphology, and pappus structure, the combination Richteria tianschanica (Krasch.) Sonboli & Oberpr. is proposed based on the basionym Pyrethrum tianschanicum Krasch.     

Phylogenetic position of the coral symbiont Ostreobium (Ulvophyceae) inferred from chloroplast genome data

The green algal genus Ostreobium is an important symbiont of corals, playing roles in reef decalcification and providing photosynthates to the coral during bleaching events. A chloroplast genome of a cultured strain of Ostreobium was available, but low taxon sampling and Ostreobium's early‐branching nature left doubt about its phylogenetic position. Here, we generate and describe chloroplast genomes from four Ostreobium strains as well as Avrainvillea mazei and Neomeris sp., strategically sampled early‐branching lineages in the Bryopsidales and Dasycladales respectively. At 80,584 bp, the chloroplast genome of Ostreobium sp. HV05042 is the most compact yet found in the Ulvophyceae. The Avrainvillea chloroplast genome is ~94 kbp and contains introns in infA and cysT that have nearly complete sequence identity except for an open reading frame (ORF) in infA that is not present in cysT. In line with other bryopsidalean species, it also contains regions with possibly bacteria‐derived ORFs. The Neomeris data did not assemble into a canonical circular chloroplast genome but a large number of contigs containing fragments of chloroplast genes and showing evidence of long introns and intergenic regions, and the Neomeris chloroplast genome size was estimated to exceed 1.87 Mb. Chloroplast phylogenomics and 18S nrDNA data showed strong support for the Ostreobium lineage being sister to the remaining Bryopsidales. There were differences in branch support when outgroups were varied, but the overall support for the placement of Ostreobium was strong. These results permitted us to validate two suborders and introduce a third, the Ostreobineae.     

Phylogenetic position of Nemertea derived from phylogenomic data

Nemertea and Platyhelminthes have traditionally been grouped together because they possess a so-called acoelomate organization, but lateral vessels and rhynchocoel of nemerteans have been regarded as coelomic cavities. Additionally, both taxa show spiral cleavage patterns prompting the placement of Nemertea as sister to coelomate Protostomia, that is, either to Neotrochozoa (Mollusca and Annelida) or to Teloblastica (Neotrochozoa plus Arthropoda). Some workers maintain a sister group relationship of Nemertea and Platyhelminthes as Parenchymia because of an assumed homology of G?tte's and Müller's larvae of polyclad Platyhelminthes and the pilidium larvae of heteronemerteans. So far, molecular data were only able to significantly reject a sister group relationship to Teloblastica. Although phylogenomic data are available for Platyhelminthes, Annelida, Mollusca, and Arthropoda, they are lacking for Nemertea. Herein, we present the first analysis specifically addressing nemertean phylogenetic position using phylogenomic data. More specifically, we collected expressed sequence tag data from Lineus viridis (O.F. Müller, 1774) and combined it with available data to produce a data set of 9,377 amino acid positions from 60 ribosomal proteins. Maximum likelihood analyses and Bayesian inferences place Nemertea in a clade together with Annelida and Mollusca. Furthermore, hypothesis testing significantly rejected a sister group relationship to either Platyhelminthes or Teloblastica. The Coelomata hypothesis, which groups coelomate taxa together to the exclusion of acoelomate and pseudocoelomate taxa, is not congruent with our results. Thus, the supposed acoelomate organization evolved independently in Nemertea and Platyhelminthes. In Nemertea, evolution of acoely is most likely due to a secondary reduction of the coelom as it is found in certain species of Mollusca and Annelida. Though looking very similar, the G?tte's and Müller's larvae of polyclad Platyhelminthes are not homologous to the pilidium larvae of heteronemerteans. Finally, the convergent evolution of segmentation in Annelida and Arthropoda is further substantiated.     

A cytogenetic analysis of the genus Neodendrocoelum (Triclada,paludicola) from Lake Ohrid

Nine species of the genus Neodendrocoelum from Lake Ohrid, five from sublittoral and four from littoral regions, have a diploid chromosome complement 2n=32 and show a marked resemblance in their karyotypes, comprising four large pairs (group L), seven medium pairs (group M) and five small pairs (group S). Polymorphism has only been found in groups S. Variations in chiasma frequency in the species of this genus indicate that their meiotic systems are different. In these species the number of quadrivalent were found to be different in metaphase I. The process of speciation of this genus was manifested in the diploidization of autotetraploid species.     

Phylogenetic relationships of the genus Dugesia (Platyhelminthes, Tricladida, Paludicola)

A phylogenetic analysis was performed on the genera and subgenera within the freshwater triclad family Dugesiidae, based on 19 terminal taxa and 17 morphological characters. The phylogenetic tree proposed has length of 27 steps and consistency index of 0.66. This phylogenetic hypothesis implies that the current genus Dugesia is paraphyletic and that its subgenera Girardia, Schmidiea and Dugesia S.S. should be elevated to the rank of genus. The genera Cura, Spathula and Neppia are presumed monophyletic by default because the database was unable to provide autapomorphies for any of these genera. The genera Dugesia S.S. and Neppia share sistergroup relationship. Several characters are discussed which were previously considered to be of phylogenetic importance but were not included in the present analysis. It is emphasized that sensory organs form potentially useful set of phylogenetic characters for the Dugesiidae.     

Phylogenetic position and generic differentiation of Epithemateae (Gesneriaceae) inferred from plastid DNA sequence data

The systematic position and generic differentiation of the morphologically and geographically outstanding tribe Epithemateae (Gesneriaceae) was analyzed using the rbcL/atpB-spacer and trnL-F intron-spacer regions of chloroplast DNA. In our analysis Epithemateae forms a strongly supported monophyletic clade (bootstrap [BS] = 100%; jackknife [JK] = 100%; decay index [DI] = 12) and appears as sister to the rest of the paleotropical Gesneriaceae (= subfamily Cyrtandroideae). The paleotropical Gesneriaceae form a monophyletic group (BS = 88%; JK = 85%; DI = 3) that is sister to the neotropical Gesneriaceae (subfamily Gesnerioideae) plus Austral Gesneriaceae (subfamily Coronantheroideae) (BS = 99%; JK = 98%; DI = 10). Within Epithemateae Rhynchoglossum is sister to the remaining Epithemateae (BS = 97%; JK = 96%; DI = 12), in which Epithema is sister to a clade of two genera: Loxonia/Stauranthera (BS = 68%; JK = 64%; DI = 1), which form, together with Epithema, a sister clade (BS = 85%; JK = 83%; DI = 2) to Whytockia and Monophyllaea. While the support for Loxonia and Stauranthera is moderate, the relationship of Whytockia and Monophyllaea is very strongly supported (BS = 100%; JK = 100%; DI = 13). Apart from the somewhat surprising (but well-substantiated) isolated position of Rhynchoglossum, the results are in perfect accordance with the relationships worked out earlier on grounds of architectural and floral characters. Especially remarkable is the predicted coherence between the morphologically and geographically different genera Whytockia and Monophyllaea.     

Phylogenetic position of the Sphaeropleaceae (Chlorophyta)

 The complete 18S rRNA gene sequences of four Sphaeroplea C.A. Agardh strains (Sphaeropleales, Sphaeropleaceae), two Atractomorpha Hoffman strains (Sphaeropleales, Sphaeropleaceae) and two Ankyra Fott strains (Chlorococcales, Characiaceae) were determined and subjected to phylogenetic analyses. The analyses indicated that all these taxa belong to a monophyletic lineage (Sphaeropleaceae) and are related to a group of chlorophycean algae comprising autosporic taxa and taxa that reproduce by zoospores which are characterized by directly opposed basal bodies. The taxonomic assignment of the Sphaeropleaceae as a family within the Sphaeropleales (Chlorophyta, Chlorophyceae) is discussed. Received December 22, 2000 Accepted September 25, 2001     

Phylogenetic position of Dictyostelium inferred from multiple protein data sets

The phylogenetic position of Dictyostelium inferred from 18S rRNA data contradicts that from protein data. Protein trees always show the close affinity of Dictyostelium with animals, fungi, and plants, whereas in 18S rRNA trees the branching of Dictyostelium is placed at a position before the massive radiation of protist groups including the divergence of the three kingdoms. To settle this controversial issue and to determine the correct position of Dictyostelium, we inferred the phylogenetic relationship among Dictyostelium and the three kingdoms Animalia, Fungi, and Plantae by a maximum-likelihood method using 19 different protein data sets. It was shown at the significance level of 1 SE that the branching of Dictyostelium antedates the divergence of Animalia and Fungi, and Plantae is an outgroup of the Animalia-Fungi-Dictyostelium clade.Correspondence to: T. Miyata     

Phylogenetic position of Octosporea muscaedomesticae (Microsporidia) and its relationship to Octosporea bayeri based on small subunit rDNA analysis

Comparative phylogenetic analysis of the small subunit rDNA sequence of Octosporea muscaedomesticae (Flu, 1911) (type species) (Microsporidia) isolated from the blowfly Phormia regina (Diptera:Calliphoridae) is presented. Neighbor Joining bootstrap, Maximum Parsimony and Maximum Likelihood analyses with 38 microsporidian taxa representing five major clades of Microsporidia placed O. muscaedomesticae on a separate branch within a clade containing parasites of freshwater hosts. O. muscaedomesticae differed from Octosporea bayeri, a parasite of the microcrustacean, Daphnia magna (Cladocera:Daphniidae) by 29% demonstrating that the latter microsporidium is not closely related to the type species at the generic level, and should not be placed within the genus Octosporea, a conclusion that is further supported by morphological and developmental differences. Considering the number of disparately related hosts from which Octosporea species have been previously described based mostly on developmental and morphological characters it is likely that many will not fit the current definition of the genus, and it is possible that molecular analysis of these species will show that this genus as defined represents a polyphyletic grouping of unrelated taxa.     

Parallel evolution evidenced by molecular data in the banded-tetra (Astyanax fasciatus)

Astyanax is well known as a model for developmental biology studies, particularly with regard to Mexico's cave populations. More than 130 species of Astyanax are already known, most of which live in South America. The occurrence of cryptic species and species complexes elucidated by chromosomal and genetic studies demonstrates that the relationship between morphology and molecular evolution is quite complex within this group. In this work, we demonstrate that morphology does not follow the path of vicariant processes observed in Astyanax fasciatus populations, which separated about three million years ago, although molecular data suggests its separation in two species.     

Systematics of the Dactylorhiza euxina/incarnata/maculata polyploid complex (Orchidaceae) in Turkey: evidence from allozyme data

 Material of Dactylorhiza were sampled from 49 localities in Turkey and investigated for allozyme variation at ten loci (nine enzyme systems). Among diploids, the Anatolian D. osmanica and D. umbrosa were allozymically variable, but not distinct from each other or from D. incarnata. Dactylorhiza saccifera contained the same alleles as the European D. fuchsii. Dactylorhiza iberica and D. euxina were distinct from each other and the other diploids. On basis of allozyme patterns three distinct allotetraploid genotypes were distinguished, and each of them could be treated as a separate species. Dactylorhiza nieschalkiorum is similar to European allotetraploids, and may have arisen from hybridization between D. incarnata s.l. and D. saccifera. Dactylorhiza urvilleana may have arisen from parents related to present-day D. saccifera and D. euxina, but it also contains additional alleles that have not been found in any of the diploids investigated. A third allotetraploid known from four populations in the Ardahan and Kars provinces of north-eastern Turkey combines the allozyme patterns found in material of D. incarnata s.l. from the same area with those from D. euxina. It is here described for the first time as D. armeniaca. Received November 14, 2000 Accepted June 20, 2001