Arthropod relationships revealed by phylogenomic analysis of nuclear protein-coding sequences.

The remarkable antiquity, diversity and ecological significance of arthropods have inspired numerous attempts to resolve their deep phylogenetic history, but the results of two decades of intensive molecular phylogenetics have been mixed. The discovery that terrestrial insects (Hexapoda) are more closely related to aquatic Crustacea than to the terrestrial centipedes and millipedes (Myriapoda) was an early, if exceptional, success. More typically, analyses based on limited samples of taxa and genes have generated results that are inconsistent, weakly supported and highly sensitive to analytical conditions. Here we present strongly supported results from likelihood, Bayesian and parsimony analyses of over 41 kilobases of aligned DNA sequence from 62 single-copy nuclear protein-coding genes from 75 arthropod species. These species represent every major arthropod lineage, plus five species of tardigrades and onychophorans as outgroups. Our results strongly support Pancrustacea (Hexapoda plus Crustacea) but also strongly favour the traditional morphology-based Mandibulata (Myriapoda plus Pancrustacea) over the molecule-based Paradoxopoda (Myriapoda plus Chelicerata). In addition to Hexapoda, Pancrustacea includes three major extant lineages of 'crustaceans', each spanning a significant range of morphological disparity. These are Oligostraca (ostracods, mystacocarids, branchiurans and pentastomids), Vericrustacea (malacostracans, thecostracans, copepods and branchiopods) and Xenocarida (cephalocarids and remipedes). Finally, within Pancrustacea we identify Xenocarida as the long-sought sister group to the Hexapoda, a result confirming that 'crustaceans' are not monophyletic. These results provide a statistically well-supported phylogenetic framework for the largest animal phylum and represent a step towards ending the often-heated, century-long debate on arthropod relationships.

Resolving arthropod phylogeny: exploring phylogenetic signal within 41 kb of protein-coding nuclear gene sequence.

This study attempts to resolve relationships among and within the four basal arthropod lineages (Pancrustacea, Myriapoda, Euchelicerata, Pycnogonida) and to assess the widespread expectation that remaining phylogenetic problems will yield to increasing amounts of sequence data. Sixty-eight regions of 62 protein-coding nuclear genes (approximately 41 kilobases (kb)/taxon) were sequenced for 12 taxonomically diverse arthropod taxa and a tardigrade outgroup. Parsimony, likelihood, and Bayesian analyses of total nucleotide data generally strongly supported the monophyly of each of the basal lineages represented by more than one species. Other relationships within the Arthropoda were also supported, with support levels depending on method of analysis and inclusion/exclusion of synonymous changes. Removing third codon positions, where the assumption of base compositional homogeneity was rejected, altered the results. Removing the final class of synonymous mutations--first codon positions encoding leucine and arginine, which were also compositionally heterogeneous--yielded a data set that was consistent with a hypothesis of base compositional homogeneity. Furthermore, under such a data-exclusion regime, all 68 gene regions individually were consistent with base compositional homogeneity. Restricting likelihood analyses to nonsynonymous change recovered trees with strong support for the basal lineages but not for other groups that were variably supported with more inclusive data sets. In a further effort to increase phylogenetic signal, three types of data exploration were undertaken. (1) Individual genes were ranked by their average rate of nonsynonymous change, and three rate categories were assigned--fast, intermediate, and slow. Then, bootstrap analysis of each gene was performed separately to see which taxonomic groups received strong support. Five taxonomic groups were strongly supported independently by two or more genes, and these genes mostly belonged to the slow or intermediate categories, whereas groups supported only by a single gene region tended to be from genes of the fast category, arguing that fast genes provide a less consistent signal. (2) A sensitivity analysis was performed in which increasing numbers of genes were excluded, beginning with the fastest. The number of strongly supported nodes increased up to a point and then decreased slightly. Recovery of Hexapoda required removal of fast genes. Support for Mandibulata (Pancrustacea + Myriapoda) also increased, at times to "strong" levels, with removal of the fastest genes. (3) Concordance selection was evaluated by clustering genes according to their ability to recover Pancrustacea, Euchelicerata, or Myriapoda and analyzing the three clusters separately. All clusters of genes recovered the three concordance clades but were at times inconsistent in the relationships recovered among and within these clades, a result that indicates that the a priori concordance criteria may bias phylogenetic signal in unexpected ways. In a further attempt to increase support of taxonomic relationships, sequence data from 49 additional taxa for three slow genes (i.e., EF-1 alpha, EF-2, and Pol II) were combined with the various 13-taxon data sets. The 62-taxon analyses supported the results of the 13-taxon analyses and provided increased support for additional pancrustacean clades found in an earlier analysis including only EF-1 alpha, EF-2, and Pol II.