A modern approach to rotiferan phylogeny: combining morphological and molecular data

The phylogeny of selected members of the phylum Rotifera is examined based on analyses under parsimony direct optimization and Bayesian inference of phylogeny. Species of the higher metazoan lineages Acanthocephala, Micrognathozoa, Cycliophora, and potential outgroups are included to test rotiferan monophyly. The data include 74 morphological characters combined with DNA sequence data from four molecular loci, including the nuclear 18S rRNA, 28S rRNA, histone H3, and the mitochondrial cytochrome c oxidase subunit I. The combined molecular and total evidence analyses support the inclusion of Acanthocephala as a rotiferan ingroup, but do not support the inclusion of Micrognathozoa and Cycliophora. Within Rotifera, the monophyletic Monogononta is sister group to a clade consisting of Acanthocephala, Seisonidea, and Bdelloidea-for which we propose the name Hemirotifera. We also formally propose the inclusion of Acanthocephala within Rotifera, but maintaining the name Rotifera for the new expanded phylum. Within Monogononta, Gnesiotrocha and Ploima are also supported by the data. The relationships within Ploima remain unstable to parameter variation or to the method of phylogeny reconstruction and poorly supported, and the analyses showed that monophyly was questionable for the families Dicranophoridae, Notommatidae, and Brachionidae, and for the genus Proales. Otherwise, monophyly was generally supported for the represented ploimid families and genera.     

Interrelationships of the Gastrotricha and their place among the Metazoa inferred from 18S rRNA genes

The phylum Gastrotricha includes about 700 species. They are small worm‐like organisms abundant among marine and freshwater meiobenthos. In spite of their ubiquity, diversity and relative abundance, phylogenetic relationships of these animals remain enigmatic due to the conflicting results of morphological and molecular cladistic analyses. Also unclear are the alliances within the phylum. In order to best estimate the position of Gastrotricha among the Metazoa and to shed some light on the ingroup phylogenetic relationships, small subunit (SSU) ribosomal DNA (rDNA) from 15 species of Chaetonotida (eight genera) and 28 species of Macrodasyida (26 genera) were included in an alignment of 50 metazoan taxa representing 26 phyla. Of the gastrotrich SSU rDNA sequences, eight are new and, along with published sequences represent eight families, including the five marine most speciose. Gastrotricha were resolved within a monophyletic Lophotrochozoa as part of a clade including Micrognathozoa, Rotifera and Cycliophora. The Gnathostomulida were sister to this clade. Nodal support was low for all of these relationships except the grouping of the Micrognathozoa, Rotifera and Cycliophora. Bayesian inference resolved the Gastrotricha as monophyletic with weak nodal support; the Macrodasyida were resolved as paraphyletic with many basal nodes poorly supported. Within the Chaetonotida, the monotypic Multitubulatina Neodasys was found in alliance with the macrodasyidan Urodasys while all the Paucitubulatina were found to form a single, well‐supported clade, with Musellifer as the most basal member. Among the more densely sampled Macrodasyida the Lepidodasyidae and Macrodasyidae were each found to be polyphyletic while monophyly was well supported for the Turbanellidae and Thaumastodermatidae. The congruence of our results with those of the cladistic analysis based on morphological traits provides confidence about the value of each dataset, and calls for widening of the research to include additional taxa of particular phylogenetic significance such as the Dactylopodolidae, Diuronotus, Heteroxenotrichula and Draculiciteria. The study highlights the problems in working with small species, the need for voucher specimens and the confused taxonomic status and membership of various gastrotrich families.     

The ultrastructure of the mastax of Filinia longiseta (Flosculariaceae, Rotifera): Informational value of the trophi structure and mastax musculature

The study contributes to the discussion of mastax evolution within Rotifera by giving an insight into the ultrastructure of the mastax in the rotifer species Filinia longiseta (Flosculariacea) and additionally into the bdelloid rotifer species Adineta vaga and Zelinkiella synaptae. The existence of cuticularized jaw elements (trophi) in the mastax, a muscular pharynx, is one of the defining rotiferan characters and the basis on which the monophyletic taxon Gnathifera Ahlrichs 1995a, comprising Rotifera, Gnathostomulida, Micrognathozoa and Acanthocephala, was erected. By means of SEM observations of the trophi and ultrathin serial sections (TEM) of the mastax, the internal and external organization of the jaw elements of F. longiseta is reconstructed. TEM sections of the incus of Filinia demonstrate that the fulcrum and the rami are built up by multitudes of tiny cuticular tubes. While tubular substructures in the rotiferan fulcrum have been described previously, distinct cuticular tubes as a substructure of the ramus have only been described for species belonging to the taxa Seisonidea and Bdelloidea so far ( [Koehler and Hayes, 1969] and [Ahlrichs, 1995b]). By comparing the appearance and arrangement of the cuticular tubes in the rami of F. longiseta to those found in species of Seisonidea and Bdelloidea, a higher degree of resemblance between the structures in F. longiseta and Bdelloidea can be reported. The occurrence of the ramus substructures in species of Seisonidea (Paraseison annulatus and Seison nebaliae) is given consideration to represent an intermediate between the ramus substructure of Bdelloidea/Flosculariacea and Ploima. Additionally, the mastax musculature of F. longiseta, being associated with the trophi, is described: A total of seven muscles are found that directly insert the jaw elements or are indirectly associated with them via muscle-to-muscle connections.     

Micrognathozoa: a new class with complicated jaws like those of Rotifera and Gnathostomulida

A new microscopic aschelminth-like animal, Limnognathia maerski nov. gen. et sp., is described from a cold spring at Disko Island, West Greenland, and assigned to Micrognathozoa nov. class. It has a complex of jaws in its pharynx, and the ultrastructure of the main jaws is similar to that of the jaws of advanced scleroperalian gnathostomulids. However, other jaw elements appear also to have characteristics of the trophi of Rotifera. Jaw-like structures are found in other protostome taxa as well-for instance, in proboscises of kalyptorhynch platyhelminths, in dorvilleid polychaetes and aplacophoran mollusks-but studies of their ultrastructure show that none of these jaws is homologous with jaws found in Gnathostomulida, Rotifera, and Micrognathozoa. The latter three groups have recently been joined into the monophylum Gnathifera Ahlrichs, 1995, an interpretation supported by the presence of jaw elements with cuticular rods with osmiophilic cores in all three groups. Such tubular structures are found in the fulcrum of all Rotifera and in several cuticular sclerites of both Gnathostomulida and Micrognathozoa. The gross morphology of the pharyngeal apparatus is similar in the three groups. It consists of a ventral pharyngeal bulb and a dorsal pharyngeal lumen. The absence of pharyngeal ciliation cannot be used as an autapomorphy in the ground pattern of the Gnathifera because the Micrognathozoa has the plesiomorphic alternative with a ciliated pharyngeal epithelium. The body of Limnognathia maerski nov. gen. et sp. consists of a head, thorax, and abdomen. The dorsal and lateral epidermis have plates formed by an intracellular matrix, as in Rotifera and Acanthocephala; however, the epidermis is not syncytial. The ventral epidermis lacks internal plates, but has a cuticular oral plate without ciliary structures. Two ventral rows of multiciliated cells form a locomotory organ. These ciliated cells resemble the ciliophores present in some interstitial annelids. An adhesive ciliated pad is located ventrally close to a caudal plate. As in many marine interstitial animals-e.g., gnathostomulids, gastrotrichs, and polychaetes-a special form of tactile bristles or sensoria is found on the body. Two pairs of protonephridia with unicellular terminal cells are found in the trunk; this unicellular condition may be the plesiomorphic condition in Bilateria. Only specimens with the female reproductive system have been found, indicating that all adult animals are parthenogenetic females. We suggest that 1) jaws of Gnathostomulida, Rotifera, and the new taxon, Micrognathozoa, are homologous structures; 2) Rotifera (including Acanthocephala) and the new group might be sister groups, while Gnathostomulida could be the sister-group to this assemblage; and 3) the similarities to certain gastrotrichs and interstitial polychaetes are convergent.     

Ontogeny of the jaws of monogonont rotifers: the malleate trophi of Rhinoglena and Proalides (Ploima,Epiphanidae)

Information on the embryonic development of the malleate trophi in Epiphanidae (Rotifera, Monogononta, Ploima) is presented, based on scanning electron microscopy observations in Rhinoglena fertoeensis, R. frontalis, R. kutikovae, R. tokioensis, and Proalides tentaculatus, to contribute to the understanding of this structure of high evolutionary and functional relevance in Rotifera. The first observable and distinctly sclerotized structures were a double row of median transversal sclerites along the longitudinal axis, wherein the future unci, rostellar scleropili, cristae rami, and basal apophyses became recognizable. Fulcrum and manubria arose subsequently; the fulcrum sclerites were longitudinally ordered in a double layer. The rami chambers developed last as lamellar structures. Unci appeared as separate thin, elongate elements, the primary uncini, developing to uncus plates by transversal growth and apposition of sclerite material on the shafts of the uncini. The heads of the uncini showed their greatest development after fusion of their shafts into uncus plates. The interjacent spaces between the heads functioned as a mold, organizing bundles of sclerites which developed into the uniseriate, zigzag‐shaped cristae rami. The fulcrum attained its definite shape by elongation of the double layer of rod‐shaped sclerites into appressed sclerofibrillae. Manubria became visible as a proximal ridge of sclerites, whereupon a triangular lamella composed of crisscross‐oriented sclerites developed distally, growing out to the manubrial chambers. Ramus chambers originated from two longitudinal amorphous lamellae incorporating the median rami sclerites and closing from distal to proximal; subbasal chambers were formed before the basal chambers.     

Investigations into the phylogenetic position of Micrognathozoa using four molecular loci

Micrognathozoa is the most recently discovered higher metazoan lineage. The sole known species of the group, Limnognathia maerski, was originally reported from running freshwater in Disko Island (Greenland), and has recently been recorded from the subantarctic region. Because of the presence of a particular type of jaws formed of special cuticularized rods, similar to those of gnathostomulids and rotifers, the three metazoan lineages were considered closely related, and assigned to the clade Gnathifera. A phylogenetic comparison of four molecular loci for Limnognathia maerski and other newly generated sequences of mainly acoelomate animals showed that Micrognathozoa may constitute an independent lineage from those of Gnathostomulida and Rotifera. However, the exact position of Micrognathozoa could not be determined due to the lack of support for any given relationships and due to the lack of stability in the position of Limnognathia maerski under analysis of different loci and of different parameter sets for sequence comparison. Nuclear loci tend to place Micrognathozoa with the syndermatan/cycliophoran taxa, but the addition of the mitochondrial gene cytochrome c oxidase subunit I favors a relationship of Micrognathozoa to Entoprocta.     

Molecular evidence for Acanthocephala as a subtaxon of Rotifera

Rotifers are free-living animals usually smaller than 1 mm that possess a characteristic wheel organ. Acanthocephalans (thorny-headed worms) are larger endoparasitic animals that use vertebrates and arthropods to complete their life cycle. The taxa Acanthocephala and Rotifera are considered separate phyla, often within the taxon Aschelminthes. We have reexamined the relationship between Rotifera and Acanthocephala using 18S rRNA gene sequences. Our results conclusively show that Acanthocephala is the sister group of the rotifer class Bdelloidea. Rotifera was nonmonophyletic in all molecular analyses, which supports the hypothesis that the Acanthocephala represent a taxon within the phylum Rotifera and not a separate phylum. These results agree with a previous cladistic study of morphological characters. Correspondence to: J.R. Garey     

Study of the Trophi of <Emphasis Type="Italic">Testudinella</Emphasis> Bory de St. Vincent and <Emphasis Type="Italic">Pompholyx</Emphasis> Gosse (Rotifera: Testudinellidae) by Scanning Electron Microscopy

The fine morphology of the trophi of Pompholyx sulcata and nine species of Testudinella (Rotifera, Monogononta, Flosculariacea, Testudinellidae) was studied by scanning electron microscopy. The number of unci teeth and arched rami scleropili, and the shape of the major unci teeth and fulcrum are considered to be reliable additional characters for identification.     

Further structures in the jaw apparatus of Limnognathia maerski (Micrognathozoa), with notes on the phylogeny of the Gnathifera

The jaws of Limnognathia maerski, Micrognathozoa, were investigated with light- and scanning electron microscopy. The study yielded several new structures and sclerites, including the ventral part of main jaw, the pharyngeal lamellae, the manus, the dorsal and ventral fibularium teeth, and a reinterpretation of the fibularium compartmentalization. Furthermore, it was shown that several jaw elements are composed of densely packed rods. Comparison with Rotifera and Gnathostomulida suggested that the micrognathozoan main jaw is homologous with the rotifer incus and the gnathostomulid articularium and that the pseudophalangids (the ventral jaws) and their associated sclerites correspond to the rotifer mallei. These results imply that Micrognathozoa is more closely related to Rotifera than to Gnathostomulida.     

An introduction to loricifera, cycliophora, and micrognathozoa

Loriciferans, cycliophorans and micrognathozoans are amongstthe latest groups of animals to be discovered. Other than allbeing microscopic, they have very different body plans and arenot closely related. Loriciferans were originally assigned tothe Aschelminthes. However, both new molecular and ultrastructuralresearches have shown that Aschelminthes consist of two unrelatedgroups, Cycloneuralia and Gnathifera. Cycloneuralia may be includedin the Ecdysozoa, including all molting invertebrates, and Gnathiferaare more closely related to Platyhelminthes. The phylum Loriciferashares many apomorphic characters (e.g., scalids on the introvert)with both Priapulida and Kinorhyncha, and can be included inthe taxon Scalidophora, a subgroup of Cycloneuralia. Cycliophorawas originally allied to the Entoprocta and Ectoprocta (Bryozoa)based on ultrastructual research. Subsequent molecular datashow they may be related to Rotifera and Acanthocephala, withinthe taxon Gnathifera. The phylogenetic position of Cycliophorais therefore not settled, and more ultrastructural and moleculardata are needed. Micrognathozoa is the most recent major groupof animals to be described. They show strong affinities withboth Rotifera and Gnathostomulida (within the taxon Gnathifera),especially in the fine structure of the pharyngeal apparatus,where the jaw elements have cuticular rods with osmiophiliccores. Furthermore the micrognathozoans have two rows of multiciliatedcells that form a locomotory organ, similar to that seen insome gastrotrichs and interstitial annelids. This characteris never seen in Rotifera or in the monociliated Gnathostomulida.Rotifera and Acanthocephala always have a syncytial epidermis(Syndermata). Micrognathozoa lack this characteristic feature.Therefore, they are postulated to be placed basally in the Gnathifera,either as a sister-group to Gnathostomulida or as a sister-groupto Rotifera + Acanthocephala.     

Body musculature of Beauchampiella eudactylota (Gosse, 1886) (Rotifera: Euchlanidae) with additional new data on its trophi and overall morphology

This study presents the results of confocal laser scanning microscopy and fluorescence‐labelled phalloidin used to visualize the system of body musculature in Beauchampiella eudactylota. Moreover, the poorly known trophi of B. eudactylota are described based on scanning electron microscopy. In total, four paired longitudinal muscles (musculi longitudinales I–IV) and three circular muscles (musculi circulares I–III) were identified. Among these are the musculus longitudinalis ventralis, the musculus longitudinalis dorsalis and the musculus circumpedalis as documented in previous studies for other rotifer species. Compared to other species, B. eudactylota is characterized by the low number of lateral longitudinal muscles and the absence of some longitudinal muscles (musculi longitudinales capitum) and circular muscles (corona sphincter, musculus pars coronalis). Moreover, scanning electron microscopic data on the trophi of B. eudactylota reveal a number of striking similarities to the trophi in some species of Epiphanidae. This suggests that either (1) these similarities represent plesiomorphic characters present both in Epiphanidae and B. eudactylota or (2) they are synapomorphic features of B. eudactylota and some species of Epiphanidae, which would question the monophyly of Euchlanidae.     

On the Phylogenetic Position of Rotifera – Have We Come Any Further?

Rotifers are bilateral symmetric animals belonging to Protostomia. The ultrastructure of the rotiferan trophi suggests that they belong to the Gnathifera, and ultrastructural similarities between the integuments and spermatozoa as well as molecular evidence strongly suggest that rotifers and the parasitic acanthocephalans are closely related and form the clade Syndermata. Here we discuss the phylogenetic position of rotifers with regard to the gnathiferan groups. Originally, Gnathifera only included the hermaphroditic Gnathostomulida and the Syndermata. The synapomorphy supporting Gnathifera is the presence of pharyngeal hard parts such as jaws and trophi with similar ultrastructure. The newly discovered Micrognathozoa possesses such jaws and is a strong candidate for inclusion in Gnathifera because their cellular integument also has an apical intracytoplasmic lamina as in Syndermata. But Gnathifera might include other taxa. Potential candidates include the commensalistic Myzostomida and Cycliophora. Traditionally, Myzostomida has been included in the annelids but recent studies regard them either as sister group to the Acanthocephala or Cycliophora. Whether Cycliophora belongs to Gnathifera is still uncertain. Some analyses based on molecular data or total evidence point towards a close relationship between Cycliophora and Syndermata. Other cladistic studies using molecular data, morphological characters or total evidence suggest a sister group relationship between Cycliophora and Entoprocta. More molecular and morphological data and an improved sampling of taxa are obviously needed to elucidate the phylogenetic position of the rotifers and identify which phyla belong to Gnathifera.     

Phylogeny of the Acanthocephala based on morphological characters

Only four previous studies of relationships among acanthocephalans have included cladistic analyses, and knowledge of the phylogeny of the group has not kept pace with that of other taxa. The purpose of this study is to provide a more comprehensive analysis of the phylogenetic relationships among members of the phylum Acanthocephala using morphological characters. The most appropriate outgroups are those that share a common early cell-cleavage pattern (polar placement of centrioles), such as the Rotifera, rather than the Priapulida (meridional placement of centrioles) to provide character polarity based on common ancestry rather than a general similarity likely due to convergence of body shapes. The phylogeny of 22 species of the Acanthocephala was evaluated based on 138 binary and multistate characters derived from comparative morphological and ontogenetic studies. Three assumptions of cement gland structure were tested: (i) the plesiomorphic type of cement glands in the Rotifera, as the sister group, is undetermined; (ii) non-syncytial cement glands are plesiomorphic; and (iii) syncytial cement glands are plesiomorphic. The results were used to test an early move of Tegorhynchus pectinarius to Koronacantha and to evaluate the relationship between Tegorhynchus and Illiosentis. Analysis of the data-set for each of these assumptions of cement gland structure produced the same single most parsimonious tree topology. Using Assumptions i and ii for the cement glands, the trees were the same length (length = 404 steps, CI = 0.545, CIX = 0.517, HI = 0.455, HIX = 0.483, RI = 0.670, RC = 0.365). Using Assumption iii, the tree was three steps longer (length = 408 steps, CI = 0.539, CIX = 0.512, HI = 0.461, HIX = 0.488, RI = 0.665, RC = 0.359). The tree indicates that the Palaeacanthocephala and Eoacanthocephala both are monophyletic and are sister taxa. The members of the Archiacanthocephala are basal to the other two clades, but do not themselves form a clade. The results provide strong support for the Palaeacanthocephala and the Eoacanthocephala and the hypothesis that the Eoacanthocephala is the most primitive group is not supported. Little support for the Archiacanthocephala as a monophyletic group was provided by the analysis. Support is provided for the recognition of Tegorhynchus and Illiosentis as distinct taxa, as well as the transfer of T. pectinarius to Koronacantha.     

Phylogeny and classification of the Conochilidae (Rotifera, Monogononta, Flosculariacea)

To resolve several taxonomic problems within the family Conochilidae (Rotifera, Monogononta, Flosculariacea), we initiated a comparative study of the morphology in this and related taxa using samples collected from widely separated geographical regions. As part of this study, we paid special attention to trophal morphology using scanning electron microscopy. We also constructed and analysed a data matrix comprising 19 morphological characters of 11 taxa using cladistic methods to uncover all most-parsimonious trees. The results indicate that Conochilidae share a body form with Flosculariidae, but they possess a trophal structure which clearly differentiates them from all other Flosculariacea; thus, the diagnosis of the family Conochilidae is amended to incorporate morphological characters of the trophi. The analysis of our data matrix yielded a single, most-parsimonious tree. From the topology of that tree and our scanning electron microscopy observations, we propose the following: (1) the status of Conochilidae as a separate suborder of Flosculariacea is rejected; (2) taxonomic separation of Conochilus and Conochiloides as subgenera of Conochilus is confirmed; and (3) Lacinularia causeyae Vidrine, Mclaughlin & Willis, 1985 is reallocated to a new genus within the family Conochilide, Conochilopsis gen. nov., as Conochilopsis causeyae (Vidrine et al .) comb. nov.     

Cladistic and phenetic analysis of allozyme data for nine species of sea anemones of the family Actiniidae (Cnidaria: Anthozoa)

The aim of this study was to infer from allozyme data the phylogenetic relationships of nine species of actiniid sea anemones, and also use these data to assess the various methods (phenetic and cladistic) available for phylogenetic analysis. Starch gel electrophoresis was used to obtain genetic data from 13 gene loci. The anemone Metridium senile, from the family Metridiidae, was used as an outgroup. For the phenetic analysis a matrix of pairwise unbiased genetic distances was computed and, from this, dendrograms were produced both by the Wagner distance and the UPGMA methods. For the cladistic analyses three different approaches were used: the first was to treat the allele as a binary character; this was investigated using a Wagner parsimony algorithm. Another approach used was to consider the locus as an unordered character, using the alleles as states. Finally, we used the locus as an ordered multistate character, where mutation, fixation and elimination of each allele were treated as evolutionary novelties, and the heterozygotes were used as cues for the construction of transformation series. The trees produced by the phenetic and cladistic methods were highly congruent. This result suggests that allozymes can be used to produce phylogenetic hypotheses at higher taxonomic levels than those at which they are more usually employed. The Solé difference between the various trees was the relative positions of Bunodosoma caissarum and Bunodactis verrucosa in relation to the two species of Urticina. This difference was probably due to a high rate of anagenic change in B. verrucosa, which distorted the UPGMA dendrogram. The genera Actinia and Urticina appeared monophyletic in all of the trees produced. Also, the sea anemones with specialized column structures such as verrucae and vesicles (U.felina, U. eques, B. verrucosa, B. caissarum) formed a monophyletic cluster, a result compatible with the suggestion that these structures may have appeared only once in the evolutionary history of the Actiniidae.     

HYBRIDS AND PHYLOGENETIC SYSTEMATICS II. THE IMPACT OF HYBRIDS ON CLADISTIC ANALYSIS

I examined three aspects of the cladistic treatment of a set of 17 F₁ hybrids of known parental origin: (1) impact of hybrids on consistency index (CI) and number of most parsimonious trees (Trees), (2) placement of hybrids in cladograms, and (3) impact of hybrids on hypotheses of relationship among species. The hybrids were added singly and in randomly selected sets of two to five to a data set composed of Central American species of Aphelandra (including the parents of all hybrids). Compared to analyses with the same number of OTUs all of which were species, the analyses with hybrids yielded results with significantly higher CI. There was no difference in Trees between analyses with hybrids versus species. There was thus no evidence that hybrids would appear to be more problematic for cladistic methods than species. Accordingly, hybrids will not be readily identifiable as taxa that cause marked change in these indices. About % of the hybrids were placed as the cladistically basal members of the lineage that included the most apomorphic parent. Relatively apomorphic hybrids were placed proximate to the most derived parent (ca. 13% of hybrids). Other placements occurred more rarely. The most frequent placements of hybrids thus did not distinguish them from normal intermediate or apomorphic taxa. When analyses with hybrids yielded multiple most parsimonious trees, these were no more different from each other than were the equally parsimonious trees that resulted from analyses with species. Most analyses with one or two hybrids resulted in minor or no change in topology. When hybrids caused topological change, they frequently caused rearrangements of weakly supported portions of the cladogram that did not include their parents. When they disrupted the cladistic placement of their parents, they often caused their parents to change positions, with at least one topology bringing the parental lineages into closer proximity with the hybrid placed between them. Hybrids between parents from the two main lineages of the group caused total cladistic restructuring. In fact, the degree of relationship between a hybrid's parents (measured by both cladistic and patristic distance) was strongly correlated with CI (negatively) and with the degree of disturbance to cladistic relationships (positively). Thus, hybrids between distantly related parents resulted in cladograms with low CI and major topological changes. This study suggests that hybrids are unlikely to cause breakdown of cladistic structure unless they are between distantly related parents. However, these results also indicate that cladistics may not be specially useful in distinguishing hybrids from normal taxa. The applicability of these results to other kinds of hybrids is examined and the likely cladistic treatment of hybrids using other sources of data is discussed.     

Ultrastructure of spermatozoa in Orthalicidae (Mollusca,Gastropoda, Stylommatophora) and its systematic implications

Sperm morphology of orthalicid gastropods Clessinia pagoda, Spixia tucumanensis, Plagiodontes daedaleus (Odontostominae) and Drymaeus hygrohylaeus, D. poecilus, Bostryx stelzneri (Bulimulinae) are examined and described for the first time using transmission electron microscopy. Spermatozoa show the general characteristic of Pulmonata: an acrosomal vesicle, sperm nucleus helical, mitochondrial derivative forming a continuous sheath with paracrystalline material and coarse fibers associated with axonemal doublets. Features in the acrosomal complex and shape of the nucleus distinguish orthalicid sperms from other stylommatophoran. The acrosomal pedestal is traversed by fine striations in all species examined except in S. tucumanensis. The structure and thickness of the perinuclear sheath with a single or double layer of electron-dense material ensheathing the nuclear apex is characteristic of the group. The presence of a subnuclear ring in Drymaeus, Bostryx and Clessinia species is also reported. A data matrix of eleven species per 34 characters (16 sperm plus 18 anatomical and shell characters) from orthalicids plus other stylommatophoran and systellommatophoran representative species was constructed. Three cladistic analyses (sperm-based, anatomical-based and a combined sperm + anatomical-based) were performed to test the phylogenetic potential of sperm ultrastructure in orthalicid systematics and understand how sperm characters affect the topology and resolution of the obtained trees. Stylommatophora resulted in a monophyletic clade in the sperm-based and in the combined-character analysis. Orthalicidae is monophyletic only in the combined-character cladogram. Within Orthalicidae, Odontostominae is recovered as a monophyletic clade in all analyses, while Bulimulinae is paraphyletic in all trees except in the combined phylogeny. The present study and cladistic analyses performed support the hypothesis that characters on sperm ultrastructure are informative for stylommatophoran systematic and phylogenetic approaches, providing synapomorphies at familiar, subfamiliar and generic level.     

Evolutionary relationships of the New Caledonian heterotrophic conifer, Parasitaxus usta (Podocarpaceae), inferred from chloroplast trnL-F intron/spacer and nuclear rDNA ITS2 sequences

 The phylogenetic position of Parasitaxus (Podocarpaceae) has been inferred from a cladistic analysis of molecular characters from chloroplast and nuclear genomes including all genera of Podocarpaceae. In all 24 most parsimonious trees, based on combined datasets, Phyllocladus resided outside Podocarpaceae s. str. while Lepidothamnus was basal to the latter. Most other genera were arranged in two major clades. The evidence confirms previous studies, which have suggested a relationship between Lagarostrobos, Manoao and Parasitaxus. Parasitaxus is not directly related to its host Falcatifolium taxoides. Instead it appears to be most closely related to Manoao and Lagarostrobos. No other members of this group now occur on New Caledonia. However, if the evolution of Parasitaxus were autochthonous, a free-living member of this group must once have occurred there. An accelerated evolutionary rate of the chloroplast sequence analysed was suggested, indicating that the plant behaves like a holoparasite. Received January 4, 2002; accepted April 3, 2002 Published online: September 13, 2002     

SYSTEMATIC ANALYSIS OF SPHAEROPLEA (CHLOROPHYCEAE)1

A systematic investigation of the genus Sphaeroplea was conducted using cladistic analyses of both structural and isozyme characters for the same set of taxa. The structural data were not able to fully resolve some of the taxa while the isozyme data did produce a tree in which all nodes were supported by data. The structural characters were relatively consistent with one another, whereas the isozyme characters were much less internally consistent. Results from independent, cladistic analyses of both data sets support the concept that among those Sphaeroplea species investigated, S. fragilis Buchheim et Hoffman had an early divergence. The two data sets differed primarily in that the structural data support monophyly of the genus Sphaeroplea and the isozyme data do not. The greater relative consistency of the structural data suggests better support for trees inferred from its analysis. Furthermore, searches for character congruence between the two data sets revealed isozyme data which support monophyly of the genus Sphaeroplea, but had been overwhelmed by conflicting isozyme characters.     

Molecular data and the evolutionary history of dinoflagellates

We have sequenced small-subunit (SSU) ribosomal RNA (rRNA) genes from 16 dinoflagellates, produced phylogenetic trees of the group containing 105 taxa, and combined small- and partial large-subunit (LSU) rRNA data to produce new phylogenetic trees. We compare phylogenetic trees based on dinoflagellate rRNA and protein genes with established hypotheses of dinoflagellate evolution based on morphological data. Protein-gene trees have too few species for meaningful in-group phylogenetic analyses, but provide important insights on the phylogenetic position of dinoflagellates as a whole, on the identity of their close relatives, and on specific questions of evolutionary history. Phylogenetic trees obtained from dinoflagellate SSU rRNA genes are generally poorly resolved, but include by far the most species and some well-supported clades. Combined analyses of SSU and LSU somewhat improve support for several nodes, but are still weakly resolved. All analyses agree on the placement of dinoflagellates with ciliates and apicomplexans (=Sporozoa) in a well-supported clade, the alveolates. The closest relatives to dinokaryotic dinoflagellates appear to be apicomplexans, Perkinsus, Parvilucifera, syndinians and Oxyrrhis. The position of Noctiluca scintillans is unstable, while Blastodiniales as currently circumscribed seems polyphyletic. The same is true for Gymnodiniales: all phylogenetic trees examined (SSU and LSU-based) suggest that thecal plates have been lost repeatedly during dinoflagellate evolution. It is unclear whether any gymnodinialean clades originated before the theca. Peridiniales appear to be a paraphyletic group from which other dinoflagellate orders like Prorocentrales, Dinophysiales, most Gymnodiniales, and possibly also Gonyaulacales originated. Dinophysiales and Suessiales are strongly supported holophyletic groups, as is Gonyaulacales, although with more modest support. Prorocentrales is a monophyletic group only in some LSU-based trees. Within Gonyaulacales, molecular data broadly agree with classificatory schemes based on morphology. Implications of this taxonomic scheme for the evolution of selected dinoflagellate features (the nucleus, mitosis, flagella and photosynthesis) are discussed.