Phylogeny of marine Gregarines (Apicomplexa)--Pterospora, Lithocystis and Lankesteria--and the origin(s) of coelomic parasitism

Gregarines constitute a large group of apicomplexans with diverse modes of nutrition and locomotion that are associated with different host compartments (e.g. intestinal lumena and coelomic cavities). A broad molecular phylogenetic framework for gregarines is needed to infer the early evolutionary history of apicomplexans as a whole and the evolutionary relationships between the diverse ultrastructural and behavioral characteristics found in intestinal and coelomic gregarines. To this end, we sequenced the SSU rRNA gene from (1) Lankesteria abbotti from the intestines of two Pacific appendicularians, (2) Pterospora schizosoma from the coelom of a Pacific maldanid polychaete, (3) Pterospora floridiensis from the coelom of a Gulf Atlantic maldanid polychaete and (4) Lithocystis sp. from the coelom of a Pacific heart urchin. Molecular phylogenetic analyses including the new sequences demonstrated that several environmental and misattributed sequences are derived from gregarines. The analyses also demonstrated a clade of environmental sequences that was affiliated with gregarines, but as yet none of the constituent organisms have been described at the ultrastructural level (apicomplexan clade I). Lankesteria spp. (intestinal parasites of appendicularians) grouped closely with other marine intestinal eugregarines, particularly Lecudina tuzetae, from polychaetes. The sequences from all three coelomic gregarines branched within a larger clade of intestinal eugregarines and were similarly highly divergent. A close relationship between Pterospora schizosoma (Pacific) and Pterospora floridiensis (Gulf Atlantic) was strongly supported by the data. Lithocystis sp. was more closely related to a clade of marine intestinal gregarines consisting of Lankesteria spp. and Lecudina spp. than it was to the Pterospora clade. These data suggested that coelomic parasitism evolved more than once from different marine intestinal eugregarines, although a larger taxon sample is needed to further explore this inference.     

Phylogeny of cardinalfishes (Teleostei: Gobiiformes: Apogonidae) and the evolution of visceral bioluminescence

The cardinalfishes (Apogonidae) are a diverse clade of small, mostly reef-dwelling fishes, for which a variety of morphological data have not yielded a consistent phylogeny. We use DNA sequence to hypothesize phylogenetic relationships within Apogonidae and among apogonids and other acanthomorph families, to examine patterns of evolution including the distribution of a visceral bioluminescence system. In conformance with previous studies, Apogonidae is placed in a clade with Pempheridae, Kurtidae, Leiognathidae, and Gobioidei. The apogonid genus Pseudamia is recovered outside the remainder of the family, not as sister to the superficially similar genus Gymnapogon. Species sampled from the Caribbean and Western Atlantic (Phaeoptyx, Astrapogon, and some Apogon species) form a clade, as do the larger-bodied Glossamia and Cheilodipterus. Incidence of visceral bioluminescence is found scattered throughout the phylogeny, independently for each group in which it is present. Examination of the fine structure of the visceral bioluminescence system through histology shows that light organs exhibit a range of morphologies, with some composed of complex masses of tubules (Siphamia, Pempheris, Parapriacanthus) and others lacking tubules but containing chambers formed by folds of the visceral epithelium (Acropoma, Archamia, Jaydia, and Rhabdamia). Light organs in Siphamia, Acropoma, Pempheris and Parapriacanthus are distinct from but connected to the gut; those in Archamia, Jaydia, and Rhabdamia are simply portions of the intestinal tract, and are little differentiated from the surrounding tissues. The presence or absence of symbiotic luminescent bacteria does not correlate with light organ structure; the tubular light organs of Siphamia and chambered tubes of Acropoma house bacteria, those in Pempheridae and the other Apogonidae do not.     

Molecular Phylogeny of the Ocelloid-Bearing Dinoflagellates Erythropsidinium and Warnowia (Warnowiaceae, Dinophyceae)

ABSTRACT. Members of the family Warnowiaceae are unarmored phagotrophic dinoflagellates that possess an ocelloid. The genus Erythropsidinium (= Erythropsis ) has also developed a unique dynamic appendage, the piston, which is able to independently retract and extend for at least 2 min after the cell lyses. We provide the first small subunit ribosomal RNA gene sequences of warnowiid dinoflagellates, those of the type Erythropsidinium agile and one species of Warnowia . Phylogenetic analyses show that warnowiid dinoflagellates branch within the Gymnodinium sensu stricto group, forming a cluster separated from the Polykrikos clade and with autotrophic Pheopolykrikos beauchampii as closest relative. This reinforces their classification as unarmored dinoflagellates based on the shape of the apical groove, despite the strong ecological and ultrastructural diversity of the Gymnodinium s.s. group. Other structures, such as the ocelloid and piston, have no systematic value above the genus level.     

Phylogeny and Megasystematics of Phagotrophic Heterokonts (Kingdom Chromista)

Heterokonts are evolutionarily important as the most nutritionally diverse eukaryote supergroup and the most species-rich branch of the eukaryotic kingdom Chromista. Ancestrally photosynthetic/phagotrophic algae (mixotrophs), they include several ecologically important purely heterotrophic lineages, all grossly understudied phylogenetically and of uncertain relationships. We sequenced 18S rRNA genes from 14 phagotrophic non-photosynthetic heterokonts and a probable Ochromonas, performed phylogenetic analysis of 210–430 Heterokonta, and revised higher classification of Heterokonta and its three phyla: the predominantly photosynthetic Ochrophyta; the non-photosynthetic Pseudofungi; and Bigyra (now comprising subphyla Opalozoa, Bicoecia, Sagenista). The deepest heterokont divergence is apparently between Bigyra, as revised here, and Ochrophyta/Pseudofungi. We found a third universal heterokont signature sequence, and deduce three independent losses of ciliary hairs, several of 1-2 cilia, 10 of photosynthesis, but perhaps only two plastid losses. In Ochrophyta, heterotrophic Oikomonas is sister to the photosynthetic Chrysamoeba, whilst the abundant freshwater predator Spumella is biphyletic; neither clade is specifically related to Paraphysomonas, indicating four losses of photosynthesis by chrysomonads. Sister to Chrysomonadea (Chrysophyceae) is Picophagea cl. nov. (Picophagus, Chlamydomyxa). The diatom-parasite Pirsonia belongs in Pseudofungi. Heliozoan-like actinophryids (e.g. Actinosphaerium) are Opalozoa, not related to pedinellids within Hypogyristea cl. nov. of Ochrophyta as once thought. The zooflagellate class Bicoecea (perhaps the ancestral phenotype of Bigyra) is unexpectedly diverse and a major focus of our study. We describe four new biciliate bicoecean genera and five new species: Nerada mexicana, Labromonas fenchelii (=Pseudobodo tremulans sensu Fenchel), Boroka karpovii (=P. tremulans sensu Karpov), Anoeca atlantica and Cafeteria mylnikovii; several cultures were previously misidentified as Pseudobodo tremulans. Nerada and the uniciliate Paramonas are related to Siluania and Adriamonas; this clade (Pseudodendromonadales emend.) is probably sister to Bicosoeca. Genetically diverse Caecitellus is probably related to Anoeca, Symbiomonas and Cafeteria (collectively Anoecales emend.). Boroka is sister to Pseudodendromonadales/Bicoecales/Anoecales. Placidiales are probably divergent bicoeceans (the GenBank Placidia sequence is a basidiomycete/heterokont chimaera). Two GenBank ‘opalinid’ sequences are fungal; Pseudopirsonia is cercozoan; two previous GenBank ‘Caecitellus’ sequences are Adriamonas. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editior: Patnck J. Keeling]     

Phylogeny and evolution of Loxoconcha (Ostracoda, Crustacea) species around Japan

The pore-systems of 17 extant species of Loxoconcha around Japan were studied in order to understand their phylogeny and evolution. The phylogeny was estimated by two steps. First, the 17 species were divided into two groups, Group A (12 species) and Group B (five species) by Pore pattern Below Eye tubercle (PBE) analysis. Then, intragroup relationships were estimated by Differentiation of Distributional pattern of Pore-system (DDP) analysis. PBE analysis reveals that species of Groups A and B have on average different ecological preferences. Species of Group A, which appeared in the late Pliocene, are more diverse, have both phytal and bottom-dwelling modes of life, possess fewer pore-systems in the ventral area, and inhabit normal marine environments. Species of Group B, whose oldest fossil record is the lower Miocene, are less diverse, have only bottom-dwelling species, possess more pore-systems in the ventral area, and tend to inhabit brackish water environments. The results of this study suggest that the differences in ecology may have had an impact on the late Cenozoic diversification around Japan. The primary invasion of Group B occurred before the lower Miocene,with no subsequent diversification. Group A invaded after the late Pliocene and immediately diversified, which created the present abundance of Loxoconcha species around Japan in both species diversity and variety of modes of life.     

Molecular Phylogeny of Coral-Reef Sea Cucumbers (Holothuriidae: Aspidochirotida) Based on 16S Mitochondrial Ribosomal DNA Sequence

Members of the Holothuriidae, found globally at low to middle latitudes, are often a dominant component of Indo–West Pacific coral reefs. We present the first phylogeny of the group, using 8 species from the 5 currently recognized genera and based on approximately 540 nucleotides from a polymerase chain reaction–amplified and conserved 3′ section of 16S mitochondrial ribosomal DNA. Parsimony and likelihood analyses returned identical topologies, permitting several robust inferences to be drawn. Several points corroborated the Linnean classification. Actinopyga and Bohadschia each appear monophyletic and Pearsonothuria is sister to Bohadschia. Other aspects of our phylogeny, however, were not in accord with the taxonomy of Holothuriidae or previous speculations about the group’s evolutionary history. Most notably, the genus Holothuria appears paraphyletic. Actinopyga and Bohadschia, sometimes held to be closely related to one another because of certain morphologic similarities, are only distantly related. The morphologically distinct Labidodemas, even thought to warrant separation at the family level, is nested well within Holothuria. A maximum parsimony reconstruction of ancestral ossicle form on the phylogeny indicated that, in addition to a probable bout of elaboration in ossicle form (the modification of rods or rosettes to holothuriid-type buttons), at least 2 rounds of ossicle simplification also transpired in which buttons reverted to rods or rosettes. Cuvierian tubules, defensive organs unique to numerous members of Holothuriidae, were probably present before the initial radiation of the family, but the reconstruction is ambiguous as to their ancestral function.     

ITS-2 and 18S rRNA Gene Phylogeny of Aplysinidae (Verongida, Demospongiae)

18S ribosomal DNA and internal transcribed spacer 2 (ITS-2) full-length sequences, each of which was sequenced three times, were used to construct phylogenetic trees with alignments based on secondary structures, in order to elucidate genealogical relationships within the Aplysinidae (Verongida). The first poriferan ITS-2 secondary structures are reported. Altogether 11 Aplysina sponges and 3 additional sponges (Verongula gigantea, Aiolochroia crassa, Smenospongia aurea) from tropical and subtropical oceans were analyzed. Based on these molecular studies, S. aurea, which is currently affiliated with the Dictyoceratida, should be reclassified to the Verongida. Aplysina appears as monophyletic. A soft form of Aplysina lacunosa was separated from other Aplysina and stands at a basal position in both 18S and ITS-2 trees. Based on ITS-2 sequence information, the Aplysina sponges could be distinguished into a single Caribbean–Eastern Pacific cluster and a Mediterranean cluster. The species concept for Aplysina sponges as well as a phylogenetic history with a possibly Tethyan origin is discussed.Reviewing Editor: Dr. Martin Kreitman     

Phylogenetic inference in Odontesthes and Atherina (Teleostei: Atheriniformes) with insights into ecological adaptation

This contribution provides an insight into Atheriniformes systematics based on four mitochondrial regions: 12S rRNA, cytb, COI and control region (2794bp in total). In the Atherinopsoidei (New World silversides), comparisons among five species of Odontesthes, O. argentinensis, O. bonariensis, O. smitti, O. hatcheri and O. incisa revealed a putative marine-freshwater pairing pattern of Odontesthes species, possibly driven by sea level fluctuations of South American waters. This study represents the first data on molecular phylogeny of Odontesthes species that can be of usefulness to biodiversity conservation policies. In the Atherinoidei (Old World silversides), Atherina boyeri was corroborated as a species complex constituted by a marine form, a marine with dark spots form and a brackish form. Concretely, Odontesthes and Atherina may represent geographically replicated models to study genetic adaptation and speciation of marine species to brackish and freshwater habitats. In addition, phylogenetic analyses supported Odontesthes and Atherina as monophyletic taxa and their separation into two differentiated suborders Atherinopsoidei and Atherinoidei, respectively.     

Phylogeny and divergence times of some racerunner lizards (Lacertidae: Eremias) inferred from mitochondrial 16S rRNA gene segments

Eremias, or racerunners, is a widespread lacertid genus occurring in China, Mongolia, Korea, Central Asia, Southwest Asia and Southeast Europe. It has been through a series of taxonomic revisions, but the phylogenetic relationships among the species and subgenera remain unclear. In this study, a frequently studied region of the mitochondrial 16S rRNA was used to (i) reassess the phylogenetic relationships of some Eremias species, (ii) test if the viviparous species form a monophyletic group, and (iii) estimate divergence time among lineages using a Bayesian relaxed molecular-clock approach. The resulting phylogeny supports monophyly of Eremias sensu Szczerbak and a clade comprising Eremias, Acanthodactylus and Latastia. An earlier finding demonstrating monophyly of the subgenus Pareremias is corroborated, with Eremias argus being the sister taxon to Eremias brenchleyi. We present the first evidence that viviparous species form a monophyletic group. In addition, Eremias przewalskii is nested within Eremias multiocellata, suggesting that the latter is likely a paraphyletic species or a species complex. Eremias acutirostris and Eremias persica form a clade that is closely related to the subgenus Pareremias. However, the subgenera Aspidorhinus, Scapteira, and Rhabderemias seem not to be monophyletic, respectively. The Bayesian divergence-time estimation suggests that Eremias originated at about 9.9 million years ago (with the 95% confidence interval ranging from 7.6 to 12 Ma), and diversified from Late Miocene to Pleistocene. Specifically, the divergence time of the subgenus Pareremias was dated to about 6.3 million years ago (with the 95% confidence interval ranging from 5.3 to 8.5 Ma), which suggests that the diversification of this subgenus might be correlated with the evolution of an East Asian monsoon climate triggered by the rapid uplift of the Tibetan Plateau approximately 8 Ma.     

Phylogeny of Bonatea (Orchidaceae: Habenariinae) based on molecular and morphological data

The genus Bonatea is widely distributed throughout southern and eastern Africa. Considerable debate surrounds the generic status of Bonatea, but there have been neither previous studies of evolutionary relationships among Bonatea species, nor any tests of the monophyly of the genus in relation to its close relative Habenaria. We investigated phylogenetic relationships between Bonatea and selected Habenaria species using morphology, as well as sequences of the chloroplast gene matK and the internal transcribed spacer (ITS) regions of nuclear ribosomal DNA. A fully resolved cladogram was obtained using morphological data, but neither the ITS, matK, nor combined data sets yielded well-resolved and well-supported phylogenetic structure for Bonatea. There is poor congruence between ITS and matK data for interspecific relationships in Bonatea, whilst the morphological results are largely congruent with the ITS analysis. Relative to Habenaria, there is little sequence variation between Bonatea species, which could indicate a recent and rapid radiation of Bonatea. Although the sampled Bonatea species form a distinct clade, more extensive sampling of Habenaria would be required to establish unambiguously whether or not Bonatea is monophyletic.     

Phylogeny and bionomics of Lasius austriacus (Hymenoptera,Formicidae)

Summary Phylogenetic analysis based on sequence data of the mitochondrial COI gene confirms the species status of the recently described Lasius austriacus. The five haplotypes of L. austriacus do not cluster according to their geographic origin, indicating a recent gene flow among the populations. The molecular data corroborate the morphology based hypothesis that L. austriacus belongs to the Lasius (Lasius s.str.) brunneus group. The invasive species Lasius neglectus forms a sister taxon with L. turcicus, both next related to L. austriacus. Other phylogenetic relationships within the genus Lasius are in accordance with morphological data.First data on the bionomics of L. austriacus are discussed in context with its phylogenetic position. Based on gyne and male morphology, excavations of nests, pitfall trapping and observations in formicaries, we hypothesize that L. austriacus is a mainly hypogaeic, monogynous species with nuptial flight. These are characters of the Lasius brunneus group in general, except the polygynous-polycalic, intranidally copulating L. neglectus. Aggression tests, however, revealed non-aggressive behaviour (antennation) between separated L. austriacus populations, but pronounced interspecific aggression against L. neglectus. This confirms the species status of L. austriacus and indicates a reduced level of intraspecific aggression, similar to L. neglectus.The status of L. austriacus as a native species in Central Europe is confirmed.Received 10 February 2003; revised 24 June 2003; accepted 23 July 2003.     

Phylogeny and classification of the Old World Emberizini (Aves, Passeriformes)

The phylogeny of the avian genus Emberiza and the monotypic genera Latoucheornis, Melophus and Miliaria (collectively the Old World Emberizini), as well as representatives for the New World Emberizini, the circumpolar genera Calcarius and Plectrophenax and the four other generally recognized tribes in the subfamily Emberizinae was estimated based on the mitochondrial cytochrome b gene and introns 6-7 of the nuclear ornithine decarboxylase (ODC) gene. Our results support monophyly of the Old World Emberizini, but do not corroborate a sister relationship to the New World Emberizini. Calcarius and Plectrophenax form a clade separated from the other Emberizini. This agrees with previous studies, and we recommend the use of the name Calcariini. Latoucheornis, Melophus and Miliaria are nested within Emberiza, and we therefore propose they be synonymized with Emberiza. Emberiza is divided into four main clades, whose relative positions are uncertain, although a sister relation between a clade with six African species and one comprising the rest of the species (30, all Palearctic) is most likely. Most clades agree with traditional, morphology-based, classifications. However, four sister relationships within Emberiza, three of which involve the previously recognized Latoucheornis, Melophus and Miliaria, are unpredicted, and reveal cases of strong morphological divergence. In contrast, the plumage similarity between adult male Emberiza (formerly Latoucheornis) siemsseni and the nominate subspecies of the New World Junco hyemalis is shown to be the result of parallel evolution. A further case of parallel plumage evolution, between African and Eurasian taxa, is pointed out. Two cases of discordance between the mitochondrial and nuclear data with respect to branch lengths and genetic divergences are considered to be the result of introgressive hybridization.     

Phylogeny of the New World diploid cottons (Gossypium L., Malvaceae) based on sequences of three low-copy nuclear genes

American diploid cottons (Gossypium L., subgenus Houzingenia Fryxell) form a monophyletic group of 13 species distributed mainly in western Mexico, extending into Arizona, Baja California, and with one disjunct species each in the Galapagos Islands and Peru. Prior phylogenetic analyses based on an alcohol dehydrogenase gene (AdhA) and nuclear ribosomal DNA indicated the need for additional data from other molecular markers to resolve phylogenetic relationships within this subgenus. Toward this end, we sequenced three nuclear genes, the anonymous locus A1341, an alcohol dehydrogenase gene (AdhC), and a cellulose synthase gene (CesA1b). Independent and combined analyses resolved clades that are congruent with current taxonomy and previous phylogenies. Our analyses diagnose at least two long distance dispersal events from the Mexican mainland to Baja California, following a rapid radiation of the primary lineages early in the diversification of the subgenus. Molecular data support the proposed recognition of a new species closely related to Gossypium laxum that was recently collected in Mexico.