G. Pass

Arthropods were the first animals to conquer land and air. They encompass more than three quarters of all described living species. This extraordinary evolutionary success is based on an astoundingly wide array of highly adaptive body organizations. A lack of robustly resolved phylogenetic relationships, however, currently impedes the reliable reconstruction of the underlying evolutionary processes. Here, we show that phylogenomic data can substantially advance our understanding of arthropod evolution and resolve several conflicts among existing hypotheses. We assembled a data set of 233 taxa and 775 genes from which an optimally informative data set of 117 taxa and 129 genes was finally selected using new heuristics and compared with the unreduced data set. We included novel expressed sequence tag (EST) data for 11 species and all published phylogenomic data augmented by recently published EST data on taxonomically important arthropod taxa. This thorough sampling reduces the chance of obtaining spurious results due to stochastic effects of undersampling taxa and genes. Orthology prediction of genes, alignment masking tools, and selection of most informative genes due to a balanced taxa-gene ratio using new heuristics were established. Our optimized data set robustly resolves major arthropod relationships. We received strong support for a sister group relationship of onychophorans and euarthropods and strong support for a close association of tardigrades and cycloneuralia. Within pancrustaceans, our analyses yielded paraphyletic crustaceans and monophyletic hexapods and robustly resolved monophyletic endopterygote insects. However, our analyses also showed for few deep splits that were recently thought to be resolved, for example, the position of myriapods, a remarkable sensitivity to methods of analyses.

Remipedes are a small and enigmatic group of crustaceans, first described only 30 years ago. Analyses of both morphological and molecular data have recently suggested a close relationship between Remipedia and Hexapoda. If true, the remipedes occupy an important position in pancrustacean evolution and may be pivotal for understanding the evolutionary history of crustaceans and hexapods. However, it is important to test this hypothesis using new data and new types of analytical approaches. Here, we assembled a phylogenomic data set of 131 taxa, incorporating newly generated 454 expressed sequence tag (EST) data from six species of crustaceans, representing five lineages (Remipedia, Laevicaudata, Spinicaudata, Ostracoda, and Malacostraca). This data set includes all crustacean species for which EST data are available (46 species), and our largest alignment encompasses 866,479 amino acid positions and 1,886 genes. A series of phylogenomic analyses was performed to evaluate pancrustacean relationships. We significantly improved the quality of our data for predicting putative orthologous genes and for generating data subsets by matrix reduction procedures, thereby improving the signal to noise ratio in the data. Eight different data sets were constructed, representing various combinations of orthologous genes, data subsets, and taxa. Our results demonstrate that the different ways to compile an initial data set of core orthologs and the selection of data subsets by matrix reduction can have marked effects on the reconstructed phylogenetic trees. Nonetheless, all eight data sets strongly support Pancrustacea with Remipedia as the sister group to Hexapoda. This is the first time that a sister group relationship of Remipedia and Hexapoda has been inferred using a comprehensive phylogenomic data set that is based on EST data. We also show that selecting data subsets with increased overall signal can help to identify and prevent artifacts in phylogenetic analyses.

Improved methods are described for anatomical investigation of small insects and other arthropods using serial semithin sections. The specimens were dehydrated with acidified 2,2-dimethoxypropane and embedded in ERL 4206 epoxy resin under vacuum. This procedure ensures good resin impregnation of thin, long body compartments and appendages. Furthermore, it produces excellent overall preservation of the specimen and its fragile anatomical structures. This procedure saves time and gives excellent results when sectioning difficult arthropod material. A continuous recording of serial semithin sections is possible when diamond knives are used.