Molecular phylogeny and evolution of <i>Parabasalia</i> with improved taxon sampling and new protein markers of actin and elongation factor-1α

Background: Inferring the evolutionary history of phylogenetically isolated, deep-branching groups of taxa—in particular determining the root—is often extraordinarily difficult because their close relatives are unavailable as suitable outgroups. One of these taxonomic groups is the phylum Parabasalia, which comprises morphologically diverse species of flagellated protists of ecological, medical, and evolutionary significance. Indeed, previous molecular phylogenetic analyses of members of this phylum have yielded conflicting and possibly erroneous inferences. Furthermore, many species of Parabasalia are symbionts in the gut of termites and cockroaches or parasites and therefore formidably difficult to cultivate, rendering available data insufficient. Increasing the numbers of examined taxa and informative characters (e.g., genes) is likely to produce more reliable inferences. Principal Findings: Actin and elongation factor-1a genes were identified newly from 22 species of termite-gut symbionts through careful manipulations and seven cultured species, which covered major lineages of Parabasalia. Their protein sequences were concatenated and analyzed with sequences of previously and newly identified glyceraldehyde-3-phosphate dehydrogenase and the small-subunit rRNA gene. This concatenated dataset provided more robust phylogenetic relationships among major groups of Parabasalia and a more plausible new root position than those previously reported. Conclusions/Significance: We conclude that increasing the number of sampled taxa as well as the addition of new sequences greatly improves the accuracy and robustness of the phylogenetic inference. A morphologically simple cell is likely the ancient form in Parabasalia as opposed to a cell with elaborate flagellar and cytoskeletal structures, which was defined as most basal in previous inferences. Nevertheless, the evolution of Parabasalia is complex owing to several independent multiplication and simplification events in these structures. Therefore, systematics based solely on morphology does not reflect the evolutionary history of parabasalids

Molecular Phylogeny and Evolution of Parabasalia with Improved Taxon Sampling and New Protein Markers of Actin and Elongation Factor-1α

BACKGROUND: Inferring the evolutionary history of phylogenetically isolated, deep-branching groups of taxa-in particular determining the root-is often extraordinarily difficult because their close relatives are unavailable as suitable outgroups. One of these taxonomic groups is the phylum Parabasalia, which comprises morphologically diverse species of flagellated protists of ecological, medical, and evolutionary significance. Indeed, previous molecular phylogenetic analyses of members of this phylum have yielded conflicting and possibly erroneous inferences. Furthermore, many species of Parabasalia are symbionts in the gut of termites and cockroaches or parasites and therefore formidably difficult to cultivate, rendering available data insufficient. Increasing the numbers of examined taxa and informative characters (e.g., genes) is likely to produce more reliable inferences. PRINCIPAL FINDINGS: Actin and elongation factor-1α genes were identified newly from 22 species of termite-gut symbionts through careful manipulations and seven cultured species, which covered major lineages of Parabasalia. Their protein sequences were concatenated and analyzed with sequences of previously and newly identified glyceraldehyde-3-phosphate dehydrogenase and the small-subunit rRNA gene. This concatenated dataset provided more robust phylogenetic relationships among major groups of Parabasalia and a more plausible new root position than those previously reported. CONCLUSIONS/SIGNIFICANCE: We conclude that increasing the number of sampled taxa as well as the addition of new sequences greatly improves the accuracy and robustness of the phylogenetic inference. A morphologically simple cell is likely the ancient form in Parabasalia as opposed to a cell with elaborate flagellar and cytoskeletal structures, which was defined as most basal in previous inferences. Nevertheless, the evolution of Parabasalia is complex owing to several independent multiplication and simplification events in these structures. Therefore, systematics based solely on morphology does not reflect the evolutionary history of parabasalids

Shimodaira-Hasegawa (SH) and approximately unbiased (AU) tests for alternative monophyletic relationships of parabasalian classes.

<p>Abbreviations of the classes are shown in the footnote of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone-0029938-t001" target="_blank">Table 1</a> or the legend of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone-0029938-g001" target="_blank">Figure 1</a>. Asterisks indicate that the tested monophyly was significantly different from the best ML topology at <i>P</i><0.05. Each of the monophyletic groupings of C+Tt and Tm+Tn appeared in the best ML topology.</p

Maximum likelihood phylogenetic analyses of parabasalids based on GAPDH (A), actin (B), and EF-1α (C) sequences.

<p>Unambiguously aligned protein sequences of 278 (A), 280 (B) and 274 (C) sites were used for phylogenetic inference. The species names of the parabasalids except for the genus <i>Trichonympha</i> are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone-0029938-t001" target="_blank">Table 1</a>. The GAPDH sequences of <i>Trichonympha acuta</i> and <i>Eucomonympha imla</i> and the EF-1α sequence of <i>Trichomonas tenax</i> (not shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone-0029938-t001" target="_blank">Table 1</a>) were also included in the analyses. Although the EF-1α sequences published for <i>Pentatrichomonas</i> and <i>Tritrichomonas </i><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone.0029938-Malik1" target="_blank">[57]</a> were not included in C because of their shorter sequence length, the analysis with a reducing number of sites (219 sites) demonstrated that they were related very closely to the sequences from the same taxa shown in C. The sequence accession number was indicated for each taxon. The sequences used for the concatenation are in bold. The trees were estimated in RAxML and the numbers near the nodes indicate the bootstrap values. Values below 50% are not shown. Vertical bars to the right of the trees represent the parabasalian classes according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029938#pone.0029938-Cepicka2" target="_blank">[26]</a>: C, Cristamonadea; S, Spirotrichonymphea; Tn, Trichonymphea; Tt, Tritrichomonadea; H, Hypotrichomonadea; and Tm, Trichomonadea. Scale bars correspond to 0.10 substitutions per site.</p

Phylogenetic relationship of parabasalids inferred from the concatenated dataset.

<p>The concatenated dataset comprising 278 amino acid sites of GAPDH, 280 amino acid sites of actin, 274 amino acid sites of EF-1α, and 1338 nucleotide sites of SSU rRNA gene sequences was analyzed in 28 parabasalian species. The tree was estimated in RAxML using separate models with the parameters and branch length optimized for each gene partitions individually. The supporting values (bootstrap in RAxML/Bayesian posterior probability) are indicated at the nodes. Values below 50% or 0.5 are indicated with hyphens. When the site-heterogeneous CAT model was used in each partition, the identical tree topology with similar bootstrap values was obtained (data not shown). Vertical bars to the right of the tree represent the parabasalian classes. The scale bar corresponds to 0.10 substitutions per site.</p

Molecular characterization of iron-containing superoxide dismutases in the heterotrophic dinoflagellate Crypthecodinium cohnii.

Superoxide dismutases (SODs) are a family of antioxidant enzymes that catalyse the degradation of toxic superoxide radicals in obligate and facultative aerobic organisms. Here, we report the presence of a multi-copy gene family encoding SODs in the heterotrophic dinoflagellate Crypthecodinium cohnii. All the genes identified (sod1 to sod17) have been cloned and sequenced, and shown to encode potentially functional dimeric iron-containing SOD isozymes. Our data revealed a considerable molecular heterogeneity of this enzyme in C. cohnii at both genomic and transcriptional levels. The C. cohnii SOD1, overexpressed in Escherichia coli, was active and its structure obtained by homology modeling using X-ray crystal structures of homologues exhibited the typical fold of dimeric FeSODs. Phylogenetic studies including 110 other dimeric FeSODs and closely related cambialistic dimeric SOD sequences showed that the C. cohnii SODs form a monophyletic group and have all been acquired by the same event of horizontal gene transfer. It also revealed a dichotomy within the C. cohnii SOD sequences that could be explained by an ancestral sod gene duplication followed by subsequent gene duplications within each of the two groups. Enzyme assays of SOD activity indicated the presence of two FeSOD activities in C. cohnii cell lysate whereas MnSOD and Cu/ZnSOD were not detected. These activities contrasted with the SOD repertoire previously characterized in photosynthetic dinoflagellates. To explain these differences, a hypothetical evolutionary scenario is proposed that suggests gains and losses of sod genes in dinoflagellates.Journal ArticleResearch Support, Non-U.S. Gov'tinfo:eu-repo/semantics/publishe

Proposed evolutionary relationships of parabasalids.

<p>The tree shows the relationships of the six parabasalian classes. Flagellar multiplication in a single mastigont system has occurred independently in the boxed classes. The multiplications have occurred ancestrally in two classes (marked with filled circles) and probably twice within the other class (open circle). Triangles indicate the occurrence of cytoskeletal simplification in undulating membrane (UM) and costa. See the text for details.</p

Molecular identification of <i>Pentatrichomonas hominis</i> in two patients with gastrointestinal symptoms

The trichomonad species Pentatrichomonas hominis colonises the gastrointestinal tract and is generally considered as a commensal organism in humans. However, some studies have recognised an association between diarrhoea and P. hominis infection in dogs and cats. In the present report, we have identified this species using molecular tools in two patients with gastrointestinal troubles. Our data suggest that P. hominis is a possible zoonotic species with a significant potential of transmission by water and could be the causative agent of intestinal symptoms in children

Mixed human intra- and inter-subtype infections with the parasite Blastocystis sp.

International audienceBecause of their limitations, current subtyping methods likely underestimate mixed human intra- and inter-subtype infections with Blastocystis sp. leading to erroneous data in the context of epidemiological studies. We confirmed this hypothesis by the identification of several isolates belonging to three subtypes in a patient considered at high risk of mixed infection through her lifestyle in rural area and long history of travelling

Mixed human intra- and inter-subtype infections with the parasite <i>Blastocystis</i> sp.

Because of their limitations, current subtyping methods likely underestimate mixed human intra- and inter-subtype infections with Blastocystis sp. leading to erroneous data in the context of epidemiological studies. We confirmed this hypothesis by the identification of several isolates belonging to three subtypes in a patient considered at high risk of mixed infection through her lifestyle in rural area and long history of travelling