Anke Braband

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.

Genetic aspects of mitochondrial genome evolution

Many years of extensive studies of metazoan mitochondrial genomes have established differences in gene arrangements and genetic codes as valuable phylogenetic markers. Understanding the underlying mechanisms of replication, transcription and the role of the control regions which cause e.g. different gene orders is important to assess the phylogenetic signal of such events. This review summarises and discusses, for the Metazoa, the general aspects of mitochondrial transcription and replication with respect to control regions as well as several proposed models of gene rearrangements. As whole genome sequencing projects accumulate, more and more observations about mitochondrial gene transfer to the nucleus are reported. Thus occurrence and phylogenetic aspects concerning nuclear mitochondrial-like sequences (NUMTS) is another aspect of this review.

Phylogenetic relationships within the Phyllopoda (Crustacea, Branchiopoda) based on mitochondrial and nuclear markers.

For several decades the relationships within the Branchiopoda (Anostraca + Phyllopoda) have been a matter of controversy. Interpretations of plesiomorphic or apomorphic character states are a difficult venture, in particular in the Phyllopoda. We explore the relationships within the Phyllopoda at the level of nucleotid comparisons of the two genes 12S rDNA (mitochondrial) and EF1alpha (nuclear), and at a higher molecular level based on introns found in the gene EF1alpha. Within the Phyllopoda our explorations show further evidence for a non-monophyletic Conchostraca (Spinicaudata + Cyclestherida + Laevicaudata). The monotypic Cyclestherida is more closely related to the Cladocera, both together forming the Cladoceromorpha. The Spinicaudata (Leptestheriidae, Limnadiidae, and Cyzicidae) is well supported. Spinicaudata and Cladoceromorpha form a monophylum. The position of the Laevicaudata remains unclear but we find neither support for a sister group relationship to the Spinicaudata nor for a close relationship of Laevicaudata and Cladocera. Within the Cladocera, we favour the Gymnomera concept with the monotypic Haplopoda being the sister group to the monophyletic Onychopoda. The Ctenopoda seems to be the sister group to the Gymnomera, which contradicts the common view of a more basal position of the Ctenopoda.

Mitogenomic analysis of decapod crustacean phylogeny corroborates traditional views on their relationships

Phylogenetic relationships within decapod crustaceans are highly controversial. Even recent analyses based on molecular datasets have shown largely contradictory results. Previous studies using mitochondrial genomes are promising but suffer from a poor and unbalanced taxon sampling. To fill these gaps we sequenced the (nearly) complete mitochondrial genomes of 13 decapod species: Stenopus hispidus, Polycheles typhlops, Panulirus versicolor, Scyllarides latus, Enoplometopus occidentalis, Homarus gammarus, Procambarus fallax f. virginalis, Upogebia major, Neaxius acanthus, Calocaris macandreae, Corallianassa coutierei, Cryptolithodes sitchensis, Neopetrolisthes maculatus, and add that of Dromia personata. Our new data allow for comprehensive analyses of decapod phylogeny using the mitochondrial genomes of 50 species covering all major taxa of the Decapoda. Five species of Stomatopoda and one species of Euphausiacea serve as outgroups. Most of our analyses using Maximum Likelihood (ML) and Bayesian inference (BI) of nucleotide and amino acid datasets revealed congruent topologies for higher level decapod relationships: (((((((Anomala, Brachyura), Thalassinida: Gebiidea), Thalassinida: Axiidea), (Astacidea, Polychelida), Achelata), Stenopodidea), Caridea), Dendrobranchiata). This result corroborates several traditional morphological views and adds new perspectives. In particular, the position of Polychelida is surprising. Nevertheless, some problems can be identified. In a minority of analyses the basal branching of Reptantia is not fully resolved, Thalassinida are monophyletic; Polychelida are the sister group to Achelata, and Stenopodidea are resolved as sister group to Caridea. Despite this and although some nodal supports are low in our phylogenetic trees, we think that the largely stable topology of the trees regardless of different types of analyses suggests that mitochondrial genomes show good potential to resolve the relationship within Decapoda.

PHYLOGENY OF THE HYDRODICTYACEAE (CHLOROPHYCEAE): INFERENCES FROM rDNA DATA1

The hydrodictyacean green algal lineage has been the focus of much research due to the fossil record of at least some members, their ornamented cell walls, and their distinctive reproductive strategies. The phylogeny of the family was, until recently, exclusively morphology based. This investigation examines hydrodictyacean isolates from several culture collections, focusing on sequences from ribosomal data: 18S rDNA, 26S rDNA (partial), and internal transcribed spacer (ITS)‐2 data. Results from phylogenetic analyses of independent and combined data matrices support the Hydrodictyaceae as a monophyletic lineage that includes isolates of Chlorotetraedron, Hydrodictyon, Pediastrum, Sorastrum, and Tetraedron. Phylogenetic analyses of rDNA data indicate that the three‐dimensional coenobium of Hydrodictyon is evolutionarily distinct from the three‐dimensional coenobium of Sorastrum. The more robust aspects of the ITS‐2 data corroborate the 18S+26S rDNA topology and provide a structural autapomorphy for the Hydrodictyaceae and Neochloridaceae, that is, an abridgment of helix IV in the secondary structure. The rDNA data do not support monophyly of Pediastrum but rather suggest the existence of four additional hydrodictyacean genera: Monactinus, Parapediastrum, Pseudopediastrum, and Stauridium.

Phylogeny and mitochondrial gene order variation in Lophotrochozoa in the light of new mitogenomic data from Nemertea

BackgroundThe new animal phylogeny established several taxa which were not identified by morphological analyses, most prominently the Ecdysozoa (arthropods, roundworms, priapulids and others) and Lophotrochozoa (molluscs, annelids, brachiopods and others). Lophotrochozoan interrelationships are under discussion, e.g. regarding the position of Nemertea (ribbon worms), which were discussed to be sister group to e.g. Mollusca, Brachiozoa or Platyhelminthes. Mitochondrial genomes contributed well with sequence data and gene order characters to the deep metazoan phylogeny debate.ResultsIn this study we present the first complete mitochondrial genome record for a member of the Nemertea, Lineus viridis. Except two trnP and trnT, all genes are located on the same strand. While gene order is most similar to that of the brachiopod Terebratulina retusa, sequence based analyses of mitochondrial genes place nemerteans close to molluscs, phoronids and entoprocts without clear preference for one of these taxa as sister group.ConclusionAlmost all recent analyses with large datasets show good support for a taxon comprising Annelida, Mollusca, Brachiopoda, Phoronida and Nemertea. But the relationships among these taxa vary between different studies. The analysis of gene order differences gives evidence for a multiple independent occurrence of a large inversion in the mitochondrial genome of Lophotrochozoa and a re-inversion of the same part in gastropods. We hypothesize that some regions of the genome have a higher chance for intramolecular recombination than others and gene order data have to be analysed carefully to detect convergent rearrangement events.

The systematics of a small spineless Desmodesmus species, D-costato-granulatus (Sphaeropleales, Chlorophyceae), based on ITS2 rDNA sequence analyses and cell wall morphology

Desmodesmus species taxonomy is one of the most long‐standing issues in green microalgal systematics due to problems associated with phenotypic plasticity. Whereas more recent species descriptions and identifications are mainly based on cell wall structures and the use of cultures, comparisons with molecular phylogenies are largely lacking. In this study, the phylogenetic relationships between 22 clones identified as Desmodesmus costato‐granulatus (Skuja) E. H. Hegew. were assessed using ITS2 rDNA sequence data in combination with cell wall morphology. The unrooted ITS2 phylogeny showed that the clones cluster into five groups, which also differ in their cell wall structures. Therefore, the taxon is split into five species: D. costato‐granulatus, D. elegans, D. fennicus, D. regularis, and D. ultrasquamatus. Compared with other Desmodesmus species, intraspecific sequence variation is extensive and may contain additional (pseudo)cryptic diversity. Compensatory base changes were near‐absent within the species and varied from one to 11 between species. Relationships among the species were unresolved. Despite this, they clustered together with the two other Desmodesmus species having a combination of small and large warts in a well‐supported lineage. Remarkably, ITS2 sequence variation in this lineage is as high as between all other included Desmodesmus species, even though the morphology of its members is rather uniform.

The complete mitochondrial genome of Pseudocellus pearsei (Chelicerata: Ricinulei) and a comparison of mitochondrial gene rearrangements in Arachnida.

BackgroundMitochondrial genomes are widely utilized for phylogenetic and population genetic analyses among animals. In addition to sequence data the mitochondrial gene order and RNA secondary structure data are used in phylogenetic analyses. Arachnid phylogeny is still highly debated and there is a lack of sufficient sequence data for many taxa. Ricinulei (hooded tickspiders) are a morphologically distinct clade of arachnids with uncertain phylogenetic affinities.ResultsThe first complete mitochondrial DNA genome of a member of the Ricinulei, Pseudocellus pearsei (Arachnida: Ricinulei) was sequenced using a PCR-based approach. The mitochondrial genome is a typical circular duplex DNA molecule with a size of 15,099 bp, showing the complete set of genes usually present in bilaterian mitochondrial genomes. Five tRNA genes (trnW, trnY, trnN, trnL(CUN), trnV) show different relative positions compared to other Chelicerata (e.g. Limulus polyphemus, Ixodes spp.). We propose that two events led to this derived gene order: (1) a tandem duplication followed by random deletion and (2) an independent translocation of trnN. Most of the inferred tRNA secondary structures show the common cloverleaf pattern except tRNA-Glu where the TψC-arm is missing. In phylogenetic analyses (maximum likelihood, maximum parsimony, Bayesian inference) using concatenated amino acid and nucleotide sequences of protein-coding genes the basal relationships of arachnid orders remain unresolved.ConclusionPhylogenetic analyses (ML, MP, BI) of arachnid mitochondrial genomes fail to resolve interordinal relationships of Arachnida and remain in a preliminary stage because there is still a lack of mitogenomic data from important taxa such as Opiliones and Pseudoscorpiones. Gene order varies considerably within Arachnida – only eight out of 23 species have retained the putative arthropod ground pattern. Some gene order changes are valuable characters in phylogenetic analysis of intraordinal relationships, e.g. in Acari.

The complete mitochondrial genome of the sea spider Nymphon gracile (Arthropoda: Pycnogonida).

BackgroundMitochondrial genomes form units of genetic information replicating indepentently from nuclear genomes. Sequence data (most often from protein-coding genes) and other features (gene order, RNA secondary structure) of mitochondrial genomes are often used in phylogenetic studies of metazoan animals from population to phylum level. Pycnogonids are primarily marine arthropods, often considered closely related to chelicerates (spiders, scorpions and allies). However, due to their aberrant morphology and to controversial results from molecular studies, their phylogenetic position is still under debate.ResultsThis is the first report of a complete mitochondrial genome sequence from a sea spider (Nymphon gracile, class Pycnogonida). Gene order derives from that of other arthropods so that presumably 10 single tRNA gene translocations, a translocation of the mitochondrial control region, and one large inversion affecting protein-coding genes must have happened in the lineage leading to Nymphon gracile. Some of the changes in gene order seem not to be common to all pycnogonids, as those were not found in a partial mitochondrial genome of another species, Endeis spinosa. Four transfer RNAs of Nymphon gracile show derivations from the usual cloverleaf secondary structure (truncation or loss of an arm). Initial phylogenetic analyses using mitochondrial protein-coding gene sequences placed Pycnogonida as sister group to Acari. However, this is in contrast to the majority of all other studies using nuclear genes and/or morphology and was not recovered in a second analysis where two long-branching acarid species were omitted.ConclusionExtensive gene rearrangement characterizes the mitochondrial genome of Nymphon gracile. At least some of the events leading to this derived gene order happened after the split of pycnogonid subtaxa. Nucleotide and amino acid frequencies show strong differences between chelicerate taxa, presumably biasing phylogenetic analyses. Thus the affinities between Pycnogonida and Acari (mites and ticks), as found in phylogenetic analyses using mitochondrial genes, may rather be due to long-branch attraction and independently derived nucleotide composition and amino acid frequency, than to a real sister group relationship.

The mitochondrial genome of the onychophoran Opisthopatus cinctipes (Peripatopsidae) reflects the ancestral mitochondrial gene arrangement of Panarthropoda and Ecdysozoa

The ancestral genome composition in Onychophora (velvet worms) is unknown since only a single species of Peripatidae has been studied thus far, which shows a highly derived gene order with numerous translocated genes. Due to this lack of information from Onychophora, it is difficult to infer the ancestral mitochondrial gene arrangement patterns for Panarthropoda and Ecdysozoa. Hence, we analyzed the complete mitochondrial genome of the onychophoran Opisthopatus cinctipes, a representative of Peripatopsidae. Our data show that O. cinctipes possesses a highly conserved gene order, similar to that found in various arthropods. By comparing our results to those from different outgroups, we reconstruct the ancestral gene arrangement in Panarthropoda and Ecdysozoa. Our phylogenetic analysis of protein-coding gene sequences from 60 protostome species (including outgroups) provides some support for the sister group relationship of Onychophora and Arthropoda, which was not recovered by using a single species of Peripatidae, Epiperipatus biolleyi, in a previous study. A comparison of the strand-specific bias between onychophorans, arthropods, and a priapulid suggests that the peripatid E. biolleyi is less suitable for phylogenetic analyses of Ecdysozoa using mitochondrial genomic data than the peripatopsid O. cinctipes.