Mitochondrial Genome Evolution in Annelida-A Systematic Study on Conservative and Variable Gene Orders and the Factors Influencing its Evolution.

The mitochondrial genomes of Bilateria are relatively conserved in their protein-coding, rRNA, and tRNA gene complement, but the order of these genes can range from very conserved to very variable depending on the taxon. The supposedly conserved gene order of Annelida has been used to support the placement of some taxa within Annelida. Recently, authors have cast doubts on the conserved nature of the annelid gene order. Various factors may influence gene order variability including, among others, increased substitution rates, base composition differences, structure of noncoding regions, parasitism, living in extreme habitats, short generation times, and biomineralization. However, these analyses were neither done systematically nor based on well-established reference trees. Several focused on only a few of these factors and biological factors were usually explored ad-hoc without rigorous testing or correlation analyses. Herein, we investigated the variability and evolution of the annelid gene order and the factors that potentially influenced its evolution, using a comprehensive and systematic approach. The analyses were based on 170 genomes, including 33 previously unrepresented species. Our analyses included 706 different molecular properties, 20 life-history and ecological traits, and a reference tree corresponding to recent improvements concerning the annelid tree. The results showed that the gene order with and without tRNAs is generally conserved. However, individual taxa exhibit higher degrees of variability. None of the analyzed life-history and ecological traits explained the observed variability across mitochondrial gene orders. In contrast, the combination and interaction of the best-predicting factors for substitution rate and base composition explained up to 30% of the observed variability. Accordingly, correlation analyses of different molecular properties of the mitochondrial genomes showed an intricate network of direct and indirect correlations between the different molecular factors. Hence, gene order evolution seems to be driven by molecular evolutionary aspects rather than by life history or ecology. On the other hand, variability of the gene order does not predict if a taxon is difficult to place in molecular phylogenetic reconstructions using sequence data or not. We also discuss the molecular properties of annelid mitochondrial genomes considering canonical views on gene evolution and potential reasons why the canonical views do not always fit to the observed patterns without making some adjustments. [Annelida; compositional biases; ecology; gene order; life history; macroevolution; mitochondrial genomes; substitution rates.].

nRCFV: a new, dataset-size-independent metric to quantify compositional heterogeneity in nucleotide and amino acid datasets.

MOTIVATION: Compositional heterogeneity-when the proportions of nucleotides and amino acids are not broadly similar across the dataset-is a cause of a great number of phylogenetic artefacts. Whilst a variety of methods can identify it post-hoc, few metrics exist to quantify compositional heterogeneity prior to the computationally intensive task of phylogenetic tree reconstruction. Here we assess the efficacy of one such existing, widely used, metric: Relative Composition Frequency Variability (RCFV), using both real and simulated data. RESULTS: Our results show that RCFV can be biased by sequence length, the number of taxa, and the number of possible character states within the dataset. However, we also find that missing data does not appear to have an appreciable effect on RCFV. We discuss the theory behind this, the consequences of this for the future of the usage of the RCFV value and propose a new metric, nRCFV, which accounts for these biases. Alongside this, we present a new software that calculates both RCFV and nRCFV, called nRCFV_Reader. AVAILABILITY AND IMPLEMENTATION: nRCFV has been implemented in RCFV_Reader, available at: https://github.com/JFFleming/RCFV_Reader . Both our simulation and real data are available at Datadryad: https://doi.org/10.5061/dryad.wpzgmsbpn .

Multiple paths toward repeated phenotypic evolution in the spiny-leg adaptive radiation (Tetragnatha; Hawai'i).

The repeated evolution of phenotypes provides clear evidence for the role of natural selection in driving evolutionary change. However, the evolutionary origin of repeated phenotypes can be difficult to disentangle as it can arise from a combination of factors such as gene flow, shared ancestral polymorphisms or mutation. Here, we investigate the presence of these evolutionary processes in the Hawaiian spiny-leg Tetragnatha adaptive radiation, which includes four microhabitat-specialists or ecomorphs, with different body pigmentation and size (Green, Large Brown, Maroon, and Small Brown). We investigated the evolutionary history of this radiation using 76 newly generated low-coverage, whole-genome resequenced samples, along with phylogenetic and population genomic tools. Considering the Green ecomorph as the ancestral state, our results suggest that the Green ecomorph likely re-evolved once, the Large Brown and Maroon ecomorphs evolved twice and the Small Brown evolved three times. We found that the evolution of the Maroon and Small Brown ecomorphs likely involved ancestral hybridization events, while the Green and Large Brown ecomorphs likely evolved through novel mutations, despite a high rate of incomplete lineage sorting in the dataset. Our findings demonstrate that the repeated evolution of ecomorphs in the Hawaiian spiny-leg Tetragnatha is influenced by multiple evolutionary processes.

Delimitation of cryptic species drastically reduces the geographical ranges of marine interstitial ghost-worms (Stygocapitella; Annelida, Sedentaria).

The recognition of cryptic species concealed in traditionally established species may reveal new biogeographical patterns and alter the understanding of how biodiversity is geographically distributed. This is particularly relevant for marine ecosystems where the incidence of cryptic species is high and where species distribution data are often challenging to collect and interpret. Here, we studied specimens of the 'cosmopolitan' interstitial meiofaunal annelid Stygocapitella subterranea Knöllner, 1934 (Parergodrilidae, Orbiniida), obtaining data from four coastlines in the Northern hemisphere. Using phylogenetic tools and several species-delimitation methods (haplotype networks, GMYC, bPTP, maximum likelihood, posterior probability and morphology) we describe eight new Stygocapitella species. With one exception, all species are present along a single coastline, ultimately challenging the idea that Stygocapitella subterranea has a cosmopolitan distribution. We found evidence for several oceanic transitions having occurred in the past as well as a recent translocation, potentially due to human activity. No diagnostic characters were found, and qualitative and quantitative morphological data do not allow an unequivocal differentiation of the identified cryptic species. This suggests that (i) neither traditional diagnostic features nor quantitative morphology suffice to recognise species boundaries in cryptic species complexes, such as the Stygocapitella species complex; and that (ii) the recognition and description of cryptic species is of seminal importance for biodiversity assessments, biogeography and evolutionary biology.

Molecular phylogeny, morphology, and distribution of Polygordius (Polychaeta: Polygordiidae) in the Atlantic and Mediterranean.

Low morphological diversity among interstitial taxa makes it difficult to delimit species and their geographic boundaries based solely on morphology and molecular data often reveal cryptic species. Polygordius (Annelida, Polygordiidae) have low morphological diversity, but are unusual among interstitial species in their comparatively large size due to their elongated form, high fecundity, and potential for long-distance dispersal via a planktotrophic larval stage. Polygordius species collected from 14 localities in the Northwest Atlantic, Mediterranean Sea, and Southwest Atlantic including several of the respective type localities were analysed. This study presents the first phylogeny of the genus Polygordius and combines molecular data, sequences of COI, 16S and ITS1/2 genes, and morphological data for a systematic re-evaluation focusing on Atlantic species, with an emphasis on populations from European waters. Phylogenetic analyses recovered six valid species (P. appendiculatus, P. lacteus, P. neapolitanus, P. triestinus, P. jouinae, and P. eschaturus) and their distinctness is confirmed by haplotype network analyses. Thus, molecular data supported the validity of the previously recognized morpho-species and no new species were present. P. erythrophthalmus and P. villoti are invalid species being synonymous with P. lacteus. Subtle differences in head and pygidial morphology and larval type (endolarva vs. exolarva), were useful characters for discrimination. Yet seemingly significant variation in characters among individuals in some species was not diagnostic (e.g., number of pygidial cirri). Highly similar species based on adult morphology were shown to be sister taxa occurring in allopatry. Present day distribution patterns of species are summarized in light of this study.

Phylogenomics of Lophotrochozoa with Consideration of Systematic Error.

Phylogenomic studies have improved understanding of deep metazoan phylogeny and show promise for resolving incongruences among analyses based on limited numbers of loci. One region of the animal tree that has been especially difficult to resolve, even with phylogenomic approaches, is relationships within Lophotrochozoa (the animal clade that includes molluscs, annelids, and flatworms among others). Lack of resolution in phylogenomic analyses could be due to insufficient phylogenetic signal, limitations in taxon and/or gene sampling, or systematic error. Here, we investigated why lophotrochozoan phylogeny has been such a difficult question to answer by identifying and reducing sources of systematic error. We supplemented existing data with 32 new transcriptomes spanning the diversity of Lophotrochozoa and constructed a new set of Lophotrochozoa-specific core orthologs. Of these, 638 orthologous groups (OGs) passed strict screening for paralogy using a tree-based approach. In order to reduce possible sources of systematic error, we calculated branch-length heterogeneity, evolutionary rate, percent missing data, compositional bias, and saturation for each OG and analyzed increasingly stricter subsets of only the most stringent (best) OGs for these five variables. Principal component analysis of the values for each factor examined for each OG revealed that compositional heterogeneity and average patristic distance contributed most to the variance observed along the first principal component while branch-length heterogeneity and, to a lesser extent, saturation contributed most to the variance observed along the second. Missing data did not strongly contribute to either. Additional sensitivity analyses examined effects of removing taxa with heterogeneous branch lengths, large amounts of missing data, and compositional heterogeneity. Although our analyses do not unambiguously resolve lophotrochozoan phylogeny, we advance the field by reducing the list of viable hypotheses. Moreover, our systematic approach for dissection of phylogenomic data can be applied to explore sources of incongruence and poor support in any phylogenomic data set. [Annelida; Brachiopoda; Bryozoa; Entoprocta; Mollusca; Nemertea; Phoronida; Platyzoa; Polyzoa; Spiralia; Trochozoa.].

Convergent evolution of the ladder-like ventral nerve cord in Annelida.

BACKGROUND: A median, segmented, annelid nerve cord has repeatedly been compared to the arthropod and vertebrate nerve cords and became the most used textbook representation of the annelid nervous system. Recent phylogenomic analyses, however, challenge the hypothesis that a subepidermal rope-ladder-like ventral nerve cord (VNC) composed of a paired serial chain of ganglia and somata-free connectives represents either a plesiomorphic or a typical condition in annelids. RESULTS: Using a comparative approach by combining phylogenomic analyses with morphological methods (immunohistochemistry and CLSM, histology and TEM), we compiled a comprehensive dataset to reconstruct the evolution of the annelid VNC. Our phylogenomic analyses generally support previous topologies. However, the so far hard-to-place Apistobranchidae and Psammodrilidae are now incorporated among the basally branching annelids with high support. Based on this topology we reconstruct an intraepidermal VNC as the ancestral state in Annelida. Thus, a subepidermal ladder-like nerve cord clearly represents a derived condition. CONCLUSIONS: Based on the presented data, a ladder-like appearance of the ventral nerve cord evolved repeatedly, and independently of the transition from an intraepidermal to a subepidermal cord during annelid evolution. Our investigations thereby propose an alternative set of neuroanatomical characteristics for the last common ancestor of Annelida or perhaps even Spiralia.

Phylogenomic analyses unravel annelid evolution.

Annelida, the ringed worms, is a highly diverse animal phylum that includes more than 15,000 described species and constitutes the dominant benthic macrofauna from the intertidal zone down to the deep sea. A robust annelid phylogeny would shape our understanding of animal body-plan evolution and shed light on the bilaterian ground pattern. Traditionally, Annelida has been split into two major groups: Clitellata (earthworms and leeches) and polychaetes (bristle worms), but recent evidence suggests that other taxa that were once considered to be separate phyla (Sipuncula, Echiura and Siboglinidae (also known as Pogonophora)) should be included in Annelida. However, the deep-level evolutionary relationships of Annelida are still poorly understood, and a robust reconstruction of annelid evolutionary history is needed. Here we show that phylogenomic analyses of 34 annelid taxa, using 47,953 amino acid positions, recovered a well-supported phylogeny with strong support for major splits. Our results recover chaetopterids, myzostomids and sipunculids in the basal part of the tree, although the position of Myzostomida remains uncertain owing to its long branch. The remaining taxa are split into two clades: Errantia (which includes the model annelid Platynereis), and Sedentaria (which includes Clitellata). Ancestral character trait reconstructions indicate that these clades show adaptation to either an errant or a sedentary lifestyle, with alteration of accompanying morphological traits such as peristaltic movement, parapodia and sensory perception. Finally, life history characters in Annelida seem to be phylogenetically informative.

Evolution of mitochondrial gene order in Annelida.

Annelida is a highly diverse animal group with over 21,000 described species. As part of Lophotrochozoa, the vast majority of annelids are currently classified into two groups: Errantia and Sedentaria, together forming Pleistoannelida. Besides these taxa, Sipuncula, Amphinomidae, Chaetopteridae, Oweniidae and Magelonidae can be found branching at the base of the tree. Comparisons of mitochondrial genomes have been used to investigate phylogenetic relationship within animal taxa. Complete annelid mitochondrial genomes are available for some Sedentaria and Errantia and in most cases exhibit a highly conserved gene order. Only two complete genomes have been published from the basal branching lineages and these are restricted to Sipuncula. We describe the first complete mitochondrial genome sequences for all other basal branching annelid families: Owenia fusiformis (Oweniidae), Magelona mirabilis (Magelonidae), Eurythoe complanata (Amphinomidae), Chaetopterus variopedatus and Phyllochaetopterus sp. (Chaetopteridae). The mitochondrial gene order of all these taxa is substantially different from the pattern found in Pleistoannelida. Additionally, we report the first mitochondrial genomes in Annelida that encode genes on both strands. Our findings demonstrate that the supposedly highly conserved mitochondrial gene order suggested for Annelida is restricted to Pleistoannelida, representing the ground pattern of this group. All investigated basal branching annelid taxa show a completely different arrangement of genes than observed in Pleistoannelida. The gene order of protein coding and ribosomal genes in Magelona mirabilis differs only in two transposition events from a putative lophotrochozoan ground pattern and might be the closest to an ancestral annelid pattern. The mitochondrial genomes of Myzostomida show the conserved pattern of Pleistoannelida, thereby supporting their inclusion in this taxon.

Phylogeny of Syndermata (syn. Rotifera): Mitochondrial gene order verifies epizoic Seisonidea as sister to endoparasitic Acanthocephala within monophyletic Hemirotifera.

A monophyletic origin of endoparasitic thorny-headed worms (Acanthocephala) and wheel-animals (Rotifera) is widely accepted. However, the phylogeny inside the clade, be it called Syndermata or Rotifera, has lacked validation by mitochondrial (mt) data. Herein, we present the first mt genome of the key taxon Seison and report conflicting results of phylogenetic analyses: while mt sequence-based topologies showed monophyletic Lemniscea (Bdelloidea+Acanthocephala), gene order analyses supported monophyly of Pararotatoria (Seisonidea+Acanthocephala) and Hemirotifera (Bdelloidea+Pararotatoria). Sequence-based analyses obviously suffered from substitution saturation, compositional bias, and branch length heterogeneity; however, we observed no compromising effects in gene order analyses. Moreover, gene order-based topologies were robust to changes in coding (genes vs. gene pairs, two-state vs. multistate, aligned vs. non-aligned), tree reconstruction methods, and the treatment of the two monogonont mt genomes. Thus, mt gene order verifies seisonids as sister to acanthocephalans within monophyletic Hemirotifera, while deviating results of sequence-based analyses reflect artificial signal. This conclusion implies that the complex life cycle of extant acanthocephalans evolved from a free-living state, as retained by most monogononts and bdelloids, via an epizoic state with a simple life cycle, as shown by seisonids. Hence, Acanthocephala represent a rare example where ancestral transitional stages have counterparts amongst the closest relatives.

Illuminating the base of the annelid tree using transcriptomics.

Annelida is one of three animal groups possessing segmentation and is central in considerations about the evolution of different character traits. It has even been proposed that the bilaterian ancestor resembled an annelid. However, a robust phylogeny of Annelida, especially with respect to the basal relationships, has been lacking. Our study based on transcriptomic data comprising 68,750-170,497 amino acid sites from 305 to 622 proteins resolves annelid relationships, including Chaetopteridae, Amphinomidae, Sipuncula, Oweniidae, and Magelonidae in the basal part of the tree. Myzostomida, which have been indicated to belong to the basal radiation as well, are now found deeply nested within Annelida as sister group to Errantia in most analyses. On the basis of our reconstruction of a robust annelid phylogeny, we show that the basal branching taxa include a huge variety of life styles such as tube dwelling and deposit feeding, endobenthic and burrowing, tubicolous and filter feeding, and errant and carnivorous forms. Ancestral character state reconstruction suggests that the ancestral annelid possessed a pair of either sensory or grooved palps, bicellular eyes, biramous parapodia bearing simple chaeta, and lacked nuchal organs. Because the oldest fossil of Annelida is reported for Sipuncula (520 Ma), we infer that the early diversification of annelids took place at least in the Lower Cambrian.

Platyzoan paraphyly based on phylogenomic data supports a noncoelomate ancestry of spiralia.

Based on molecular data three major clades have been recognized within Bilateria: Deuterostomia, Ecdysozoa, and Spiralia. Within Spiralia, small-sized and simply organized animals such as flatworms, gastrotrichs, and gnathostomulids have recently been grouped together as Platyzoa. However, the representation of putative platyzoans was low in the respective molecular phylogenetic studies, in terms of both, taxon number and sequence data. Furthermore, increased substitution rates in platyzoan taxa raised the possibility that monophyletic Platyzoa represents an artifact due to long-branch attraction. In order to overcome such problems, we employed a phylogenomic approach, thereby substantially increasing 1) the number of sampled species within Platyzoa and 2) species-specific sequence coverage in data sets of up to 82,162 amino acid positions. Using established and new measures (long-branch score), we disentangled phylogenetic signal from misleading effects such as long-branch attraction. In doing so, our phylogenomic analyses did not recover a monophyletic origin of platyzoan taxa that, instead, appeared paraphyletic with respect to the other spiralians. Platyhelminthes and Gastrotricha formed a monophylum, which we name Rouphozoa. To the exclusion of Gnathifera, Rouphozoa and all other spiralians represent a monophyletic group, which we name Platytrochozoa. Platyzoan paraphyly suggests that the last common ancestor of Spiralia was a simple-bodied organism lacking coelomic cavities, segmentation, and complex brain structures, and that more complex animals such as annelids evolved from such a simply organized ancestor. This conclusion contradicts alternative evolutionary scenarios proposing an annelid-like ancestor of Bilateria and Spiralia and several independent events of secondary reduction.

Elucidating the phylogenetic position of Gnathostomulida and first mitochondrial genomes of Gnathostomulida, Gastrotricha and Polycladida (Platyhelminthes).

Gnathostomulida is a taxon of small marine worms, which exclusively inhabit the interstitium. The evolution of Gnathostomulida has been discussed for decades. Originally regarded as primitive animals with affinities to flatworms, the phylogenetic position of Gnathostomulida has been debated. Given the lack of an anus a close relationship to Platyhelminthes has been maintained (i.e., Plathelminthomorpha hypothesis). Alternative hypotheses proposed Gnathostomulida as being close to Gastrotricha due to the presence of a monociliary epidermis (i.e., Monokonta/Neotrichozoa hypothesis) or to Syndermata based on the complicated jaw apparatus (i.e., Gnathifera hypothesis). Molecular analyses using only few genes were inconclusive. Recent phylogenomic studies brought some progress by placing Gnathostomulida as sister to Syndermata, but support for this relationship was low and depended on the analytical strategy. Herein we present the first data of complete or nearly complete mitochondrial genomes for two gnathostomulids (Gnathostomula paradoxa &G. armata), one gastrotrich (Lepidodermella squamata) and one polyclad flatworm (Stylochoplana maculata) to address the uncertain phylogenetic affinity of Gnathostomulida. Our analyses found Gnathostomulida as sister to Syndermata (Gnathifera hypothesis). Thorough sensitivity analyses addressing taxon instability, branch length heterogeneity (also known as long branch attraction) and base composition heterogeneity showed that the position of Gnathostomulida is consistent across the different analyses and, hence, independent of potential misleading biases. Moreover, by ameliorating these different biases nodal support values could be increased to maximum values. Thus, our data support the hypothesis that the different jaw apparatuses of Syndermata and Gnathostomulida are indeed homologous structures as proposed by the Gnathifera hypothesis.

BaCoCa--a heuristic software tool for the parallel assessment of sequence biases in hundreds of gene and taxon partitions.

BaCoCa (BAse COmposition CAlculator) is a user-friendly software that combines multiple statistical approaches (like RCFV and C value calculations) to identify biases in aligned sequence data which potentially mislead phylogenetic reconstructions. As a result of its speed and flexibility, the program provides the possibility to analyze hundreds of pre-defined gene partitions and taxon subsets in one single process run. BaCoCa is command-line driven and can be easily integrated into automatic process pipelines of phylogenomic studies. Moreover, given the tab-delimited output style the results can be easily used for further analyses in programs like Excel or statistical packages like R. A built-in option of BaCoCa is the generation of heat maps with hierarchical clustering of certain results using R. As input files BaCoCa can handle FASTA and relaxed PHYLIP, which are commonly used in phylogenomic pipelines. BaCoCa is implemented in Perl and works on Windows PCs, Macs and Linux operating systems. The executable source code as well as example test files and a detailed documentation of BaCoCa are freely available at http://software.zfmk.de.

First Chromosome-Level Genome Assembly of a Ribbon Worm from the Hoplonemertea Clade, Emplectonema gracile, and Its Structural Annotation.

Genome-wide information has so far been unavailable for ribbon worms of the clade Hoplonemertea, the most species-rich class within the phylum Nemertea. While species within Pilidiophora, the sister clade of Hoplonemertea, possess a pilidium larval stage and lack stylets on their proboscis, Hoplonemertea species have a planuliform larva and are armed with stylets employed for the injection of toxins into their prey. To further compare these developmental, physiological, and behavioral differences from a genomic perspective, the availability of a reference genome for a Hoplonemertea species is crucial. Such data will be highly useful for future investigations toward a better understanding of molecular ecology, venom evolution, and regeneration not only in Nemertea but also in other marine invertebrate phyla. To this end, we herein present the annotated chromosome-level genome assembly for Emplectonema gracile (Nemertea; Hoplonemertea; Monostilifera; Emplectonematidae), an easily collected nemertean well suited for laboratory experimentation. The genome has an assembly size of 157.9 Mb. Hi-C scaffolding yielded chromosome-level scaffolds, with a scaffold N50 of 10.0 Mb and a score of 95.1% for complete BUSCO genes found as a single copy. Annotation predicted 20,684 protein-coding genes. The high-quality reference genome reaches an Earth BioGenome standard level of 7.C.Q50.

Contextualising samples: supporting reference genomes of European biodiversity through sample and associated metadata collection.

The European Reference Genome Atlas (ERGA) consortium aims to generate a reference genome catalogue for all of Europe's eukaryotic biodiversity. The biological material underlying this mission, the specimens and their derived samples, are provided through ERGA's pan-European network. To demonstrate the community's capability and capacity to realise ERGA's ambitious mission, the ERGA Pilot project was initiated. In support of the ERGA Pilot effort to generate reference genomes for European biodiversity, the ERGA Sampling and Sample Processing committee (SSP) was formed by volunteer experts from ERGA's member base. SSP aims to aid participating researchers through (i) establishing standards for and collecting of sample/specimen metadata; (ii) prioritisation of species for genome sequencing; and (iii) development of taxon-specific collection guidelines including logistics support. SSP serves as the entry point for sample providers to the ERGA genomic resource production infrastructure and guarantees that ERGA's high-quality standards are upheld throughout sample collection and processing. With the volume of researchers, projects, consortia, and organisations with interests in genomics resources expanding, this manuscript shares important experiences and lessons learned during the development of standardised operational procedures and sample provider support. The manuscript details our experiences in incorporating the FAIR and CARE principles, species prioritisation, and workflow development, which could be useful to individuals as well as other initiatives.

New phylogenomic data support the monophyly of Lophophorata and an Ectoproct-Phoronid clade and indicate that Polyzoa and Kryptrochozoa are caused by systematic bias.

BACKGROUND: Within the complex metazoan phylogeny, the relationships of the three lophophorate lineages, ectoprocts, brachiopods and phoronids, are particularly elusive. To shed further light on this issue, we present phylogenomic analyses of 196 genes from 58 bilaterian taxa, paying particular attention to the influence of compositional heterogeneity. RESULTS: The phylogenetic analyses strongly support the monophyly of Lophophorata and a sister-group relationship between Ectoprocta and Phoronida. Our results contrast previous findings based on rDNA sequences and phylogenomic datasets which supported monophyletic Polyzoa (= Bryozoa sensu lato) including Ectoprocta, Entoprocta and Cycliophora, Brachiozoa including Brachiopoda and Phoronida as well as Kryptrochozoa including Brachiopoda, Phoronida and Nemertea, thus rendering Lophophorata polyphyletic. Our attempts to identify the causes for the conflicting results revealed that Polyzoa, Brachiozoa and Kryptrochozoa are supported by character subsets with deviating amino acid compositions, whereas there is no indication for compositional heterogeneity in the character subsets supporting the monophyly of Lophophorata. CONCLUSION: Our results indicate that the support for Polyzoa, Brachiozoa and Kryptrochozoa gathered so far is likely an artifact caused by compositional bias. The monophyly of Lophophorata implies that the horseshoe-shaped mesosomal lophophore, the tentacular feeding apparatus of ectoprocts, phoronids and brachiopods is, indeed, a synapomorphy of the lophophorate lineages. The same may apply to radial cleavage. However, among phoronids also spiral cleavage is known. This suggests that the cleavage pattern is highly plastic and has changed several times within lophophorates. The sister group relationship of ectoprocts and phoronids is in accordance with the interpretation of the eversion of a ventral invagination at the beginning of metamorphosis as a common derived feature of these taxa.

The impact of mitochondrial genome analyses on the understanding of deuterostome phylogeny.

Deuterostomia, one of the three major lineages of Bilateria, comprises many well-known animals such as vertebrates, sea squirts, sea stars and sea urchins. Whereas monophyly of Deuterostomia and several subtaxa is well supported, the relationships of these to each other and, hence, deuterostome relationships are still uncertain. To address these issues in deuterostome phylogeny we analyzed datasets comprising more than 300 complete deuterostome mitochondrial genomes. Based on sequence information, the results revealed support for several relationships such as a basal position of Xenoturbella within Deuterostomia or for taxa like Craniota or Ambulacraria, but yielded also problems in some taxa, e.g. Tunicata, Pterobranchia and Ophiuroidea, due to long-branch artifacts. However, within tunicates the relationships are well supported. Variation in the genetic code was also informative and, e.g., supported the taxon Ambulacraria including Pterobranchia.

Mitochondrial genomes to the rescue--Diurodrilidae in the myzostomid trap.

Diurodrilidae is a taxon of Lophotrochozoa comprising about six, exclusively interstitial species, which are up to 500µm long and dorsoventrally flattened. Traditionally, Diurodrilidae had been regarded as an annelid family. However, recently Diurodrilidae had been excluded from Annelida and been placed in closer relationship to platyzoan taxa based on both morphological and nuclear rRNA data. Since both, Diurodrilidae and platyzoan taxa, exhibit long branches in the molecular analyses, the close relationship might be due to a long branch attraction artifact. The annelid taxon Myzostomida had been trapped in a similar long branch attraction artifact with platyzoan taxa using nuclear rRNA data, but determination of the nearly complete mitochondrial genome of myzostomids revealed their annelid affinity. Therefore, we determined the nearly complete mitochondrial genome of Diurodrilus subterraneus as well as new nuclear rRNA data for D. subterraneus and some platyzoan taxa. All our analyses of nuclear rRNA and mitochondrial sequence and gene order data presented herein clearly place Diurodrilidae within Annelida and with strong nodal support values in some analyses. Therefore, the previously suggested exclusion of Diurodrilidae from Annelida and its close relationship with platyzoan taxa can be attributed to a long branch artifact. Morphological data do not unambiguously support a platyzoan affinity of Diurodrilidae, but instead would also be in line with a progenetic origin of Diurodrilidae within Annelida.

A comprehensive analysis of bilaterian mitochondrial genomes and phylogeny.

About 2800 mitochondrial genomes of Metazoa are present in NCBI RefSeq today, two thirds belonging to vertebrates. Metazoan phylogeny was recently challenged by large scale EST approaches (phylogenomics), stabilizing classical nodes while simultaneously supporting new sister group hypotheses. The use of mitochondrial data in deep phylogeny analyses was often criticized because of high substitution rates on nucleotides, large differences in amino acid substitution rate between taxa, and biases in nucleotide frequencies. Nevertheless, mitochondrial genome data might still be promising as it allows for a larger taxon sampling, while presenting a smaller amount of sequence information. We present the most comprehensive analysis of bilaterian relationships based on mitochondrial genome data. The analyzed data set comprises more than 650 mitochondrial genomes that have been chosen to represent a profound sample of the phylogenetic as well as sequence diversity. The results are based on high quality amino acid alignments obtained from a complete reannotation of the mitogenomic sequences from NCBI RefSeq database. However, the results failed to give support for many otherwise undisputed high-ranking taxa, like Mollusca, Hexapoda, Arthropoda, and suffer from extreme long branches of Nematoda, Platyhelminthes, and some other taxa. In order to identify the sources of misleading phylogenetic signals, we discuss several problems associated with mitochondrial genome data sets, e.g. the nucleotide and amino acid landscapes and a strong correlation of gene rearrangements with long branches.