Critical analysis of the topology and rooting of the parabasalian 16S rRNA tree

The morphological classification of the protozoan phylum Parabasala is not in absolute agreement with the 16S rRNA phylogeny. However, there are strong indications that tree-construction artifacts play a considerable role in the shaping of the 16S rRNA tree. We have performed rigorous analyses designed to minimize such artifacts using the slow-fast and taxa-exclusion methods. The analyses, which included new sequences from the genera Monocercomonas and Hexamastix, in most respects confirmed the previously suggested tree topology and polyphyly of Hypermastigida and Monocercomonadidae but detected one artificial cluster of long branches (Trichonymphidae, Pseudotrichonymphidae, Hexamastix, and Tricercomitus). They also indicated that the rooting of the phylum on the trichonymphid branch is probably wrong and that reliable rooting on the basis of current data is likely impossible. We discuss the tree topology in the view of anagenesis of cytoskeletal and motility organelles and suggest that a robust taxonomic revision requires extensive analysis of other gene sequences.     

Phylogenetic identification of hypermastigotes, Pseudotrichonympha, Spirotrichonympha, Holomastigotoides, and parabasalian symbionts in the hindgut of termites

The phylogenetic diversity of parabasalian flagellates was examined based on the sequences of small subunit ribosomal RNA genes amplified directly from the mixed population of flagellates in the hindgut of lower termites. In total, 33 representative sequences of parabasalids were recovered from eight termite species. Fluorescent-labeled oligonucleotide probes specific for certain sequences were designed and used for the in situ identification of parabasalian species by whole-cell hybridization. The hypermastigotes, Pseudotrichonympha grassii, Spirotrichonympha leidyi, and Holomastigotoides mirabile in the hindgut of Coptotermes formosanus, and Spirotrichonympha sp. and Trichonympha spp. in Hodotermopsis sjoestedti were identified. In the phylogenetic tree constructed, the sequences from the termites were dispersed within the groups of known members of parabasalids, reflecting the presence of diverse parabasalids in the hindgut of termites. There were three paraphyletic lineages of hypermastigotes represented by Pseudotrichonympha, Trichonympha, and Spirotrichonympha, in agreement with the morphology-based taxonomic groups. The analysis of the tree-root suggested that the Pseudotrichonympha group is the most probable ancient lineage of parabasalids and that the Trichonympha group is the secondly deep-branching lineage. The Spirotrichonympha group and the Trichomonadida may have emerged later.