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Phylogeny of parasitic parabasalia and free-living relatives inferred from conventional markers vs. Rpb1, a single-copy gene.

Malik SB, Brochu CD, Bilic I, Yuan J, Hess M, Logsdon JM, Carlton JM - PLoS ONE (2011)

Bottom Line: As a result, genetic studies of Parabasalia lag behind other organisms.These results are consistent with prior analyses of rDNA and GAPDH sequences and ultrastructural data.These findings, together with the relative ease of Rpb1 isolation, make it an attractive tool for evaluating more extensive relationships within Parabasalia.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America. sbmalik@dal.ca

ABSTRACT

Background: Parabasalia are single-celled eukaryotes (protists) that are mainly comprised of endosymbionts of termites and wood roaches, intestinal commensals, human or veterinary parasites, and free-living species. Phylogenetic comparisons of parabasalids are typically based upon morphological characters and 18S ribosomal RNA gene sequence data (rDNA), while biochemical or molecular studies of parabasalids are limited to a few axenically cultivable parasites. These previous analyses and other studies based on PCR amplification of duplicated protein-coding genes are unable to fully resolve the evolutionary relationships of parabasalids. As a result, genetic studies of Parabasalia lag behind other organisms.

Principal findings: Comparing parabasalid EF1α, α-tubulin, enolase and MDH protein-coding genes with information from the Trichomonas vaginalis genome reveals difficulty in resolving the history of species or isolates apart from duplicated genes. A conserved single-copy gene encodes the largest subunit of RNA polymerase II (Rpb1) in T. vaginalis and other eukaryotes. Here we directly sequenced Rpb1 degenerate PCR products from 10 parabasalid genera, including several T. vaginalis isolates and avian isolates, and compared these data by phylogenetic analyses. Rpb1 genes from parabasalids, diplomonads, Parabodo, Diplonema and Percolomonas were all intronless, unlike intron-rich homologs in Naegleria, Jakoba and Malawimonas.

Conclusions/significance: The phylogeny of Rpb1 from parasitic and free-living parabasalids, and conserved Rpb1 insertions, support Trichomonadea, Tritrichomonadea, and Hypotrichomonadea as monophyletic groups. These results are consistent with prior analyses of rDNA and GAPDH sequences and ultrastructural data. The Rpb1 phylogenetic tree also resolves species- and isolate-level relationships. These findings, together with the relative ease of Rpb1 isolation, make it an attractive tool for evaluating more extensive relationships within Parabasalia.

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(A) GAPDH resolves six monophyletic parabasalid groups, but exhibits multiple nonidentical gene copies per taxon, while (B) Pms1 resolves Trichomonadea.The consensus tree topologies of the sets of best trees calculated by Bayesian inference are shown. Data generated in this study is highlighted by bold type. Scale bar represents 0.1 amino acid substitution per site. Thickened lines indicate the nodes supported by a Bayesian posterior probability of 1.00. Numbers at the nodes correspond to Bayesian posterior probabilities, followed by percent bootstrap support ≥50% given by PhyML and RAxML (1000 replicates each). The alignments are provided in Dataset S3 (GAPDH) and Dataset S4 (Pms1). (A) GAPDH. This consensus topology of the 8750 best trees calculated by Bayesian inference was constructed from 324 aligned amino acids. LnL = −7323.20, α = 1.06 (0.72<α<1.48), pI = 0.14 (0.053<pI<0.22). (B) Pms1. This consensus topology of the 9500 best trees was calculated by Bayesian inference from 538 aligned amino acids. LnL = −7126.44, α = 3.54 (2.70<α<3.98), pI = 0.040 (0.011<pI<0.071). GenBank accession numbers or Joint Genome Institute locus ID are shown at the left for each taxon.
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pone-0020774-g004: (A) GAPDH resolves six monophyletic parabasalid groups, but exhibits multiple nonidentical gene copies per taxon, while (B) Pms1 resolves Trichomonadea.The consensus tree topologies of the sets of best trees calculated by Bayesian inference are shown. Data generated in this study is highlighted by bold type. Scale bar represents 0.1 amino acid substitution per site. Thickened lines indicate the nodes supported by a Bayesian posterior probability of 1.00. Numbers at the nodes correspond to Bayesian posterior probabilities, followed by percent bootstrap support ≥50% given by PhyML and RAxML (1000 replicates each). The alignments are provided in Dataset S3 (GAPDH) and Dataset S4 (Pms1). (A) GAPDH. This consensus topology of the 8750 best trees calculated by Bayesian inference was constructed from 324 aligned amino acids. LnL = −7323.20, α = 1.06 (0.72<α<1.48), pI = 0.14 (0.053<pI<0.22). (B) Pms1. This consensus topology of the 9500 best trees was calculated by Bayesian inference from 538 aligned amino acids. LnL = −7126.44, α = 3.54 (2.70<α<3.98), pI = 0.040 (0.011<pI<0.071). GenBank accession numbers or Joint Genome Institute locus ID are shown at the left for each taxon.

Mentions: Our phylogenetic tree of Rpb1 from parabasalids and other excavates rooted with Jakoba libera (Discoba) as the outgroup is shown in Figure 3 (inferred from data in Dataset S1). Similar to recent phylogenies of multiple concatenated proteins [5], this analysis of Rpb1 also resolves Metamonada (Parabasalia, Preaxostyla (not shown) and Fornicata, represented here by the diplomonads Giardia and Spironucleus) distinct from the Discoba. Analyses of Rpb1 with and without constant sites, and with different outgroups also recover Metamonada in the majority-rule consensus topology, indicating that Discoba are at least as good as any other outgroup to Metamonada (Figure S2 and Dataset S2, and results not shown). The rooted analysis of Rpb1 in Figure 3 indicates that Hypotrichomonadea is more closely related to Tritrichomonadea than it is to Trichomonadea, a specific relationship that remains to be borne out once additional Rpb1 data is acquired from fresh isolates of uncultivable parabasalids from Cristamonadea, Spirotrichonymphea and Trichonymphea. While the relationship of Tritrichomonadea to Hypotrichomonadea shown in Figure 3 is inconsistent with results of our analyses of GAPDH and other proteins (Figures 4 and 5), it is consistent with relationships seen with some enolase and MDH paralogs (Figure 5).


Phylogeny of parasitic parabasalia and free-living relatives inferred from conventional markers vs. Rpb1, a single-copy gene.

Malik SB, Brochu CD, Bilic I, Yuan J, Hess M, Logsdon JM, Carlton JM - PLoS ONE (2011)

(A) GAPDH resolves six monophyletic parabasalid groups, but exhibits multiple nonidentical gene copies per taxon, while (B) Pms1 resolves Trichomonadea.The consensus tree topologies of the sets of best trees calculated by Bayesian inference are shown. Data generated in this study is highlighted by bold type. Scale bar represents 0.1 amino acid substitution per site. Thickened lines indicate the nodes supported by a Bayesian posterior probability of 1.00. Numbers at the nodes correspond to Bayesian posterior probabilities, followed by percent bootstrap support ≥50% given by PhyML and RAxML (1000 replicates each). The alignments are provided in Dataset S3 (GAPDH) and Dataset S4 (Pms1). (A) GAPDH. This consensus topology of the 8750 best trees calculated by Bayesian inference was constructed from 324 aligned amino acids. LnL = −7323.20, α = 1.06 (0.72<α<1.48), pI = 0.14 (0.053<pI<0.22). (B) Pms1. This consensus topology of the 9500 best trees was calculated by Bayesian inference from 538 aligned amino acids. LnL = −7126.44, α = 3.54 (2.70<α<3.98), pI = 0.040 (0.011<pI<0.071). GenBank accession numbers or Joint Genome Institute locus ID are shown at the left for each taxon.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3111441&req=5

pone-0020774-g004: (A) GAPDH resolves six monophyletic parabasalid groups, but exhibits multiple nonidentical gene copies per taxon, while (B) Pms1 resolves Trichomonadea.The consensus tree topologies of the sets of best trees calculated by Bayesian inference are shown. Data generated in this study is highlighted by bold type. Scale bar represents 0.1 amino acid substitution per site. Thickened lines indicate the nodes supported by a Bayesian posterior probability of 1.00. Numbers at the nodes correspond to Bayesian posterior probabilities, followed by percent bootstrap support ≥50% given by PhyML and RAxML (1000 replicates each). The alignments are provided in Dataset S3 (GAPDH) and Dataset S4 (Pms1). (A) GAPDH. This consensus topology of the 8750 best trees calculated by Bayesian inference was constructed from 324 aligned amino acids. LnL = −7323.20, α = 1.06 (0.72<α<1.48), pI = 0.14 (0.053<pI<0.22). (B) Pms1. This consensus topology of the 9500 best trees was calculated by Bayesian inference from 538 aligned amino acids. LnL = −7126.44, α = 3.54 (2.70<α<3.98), pI = 0.040 (0.011<pI<0.071). GenBank accession numbers or Joint Genome Institute locus ID are shown at the left for each taxon.
Mentions: Our phylogenetic tree of Rpb1 from parabasalids and other excavates rooted with Jakoba libera (Discoba) as the outgroup is shown in Figure 3 (inferred from data in Dataset S1). Similar to recent phylogenies of multiple concatenated proteins [5], this analysis of Rpb1 also resolves Metamonada (Parabasalia, Preaxostyla (not shown) and Fornicata, represented here by the diplomonads Giardia and Spironucleus) distinct from the Discoba. Analyses of Rpb1 with and without constant sites, and with different outgroups also recover Metamonada in the majority-rule consensus topology, indicating that Discoba are at least as good as any other outgroup to Metamonada (Figure S2 and Dataset S2, and results not shown). The rooted analysis of Rpb1 in Figure 3 indicates that Hypotrichomonadea is more closely related to Tritrichomonadea than it is to Trichomonadea, a specific relationship that remains to be borne out once additional Rpb1 data is acquired from fresh isolates of uncultivable parabasalids from Cristamonadea, Spirotrichonymphea and Trichonymphea. While the relationship of Tritrichomonadea to Hypotrichomonadea shown in Figure 3 is inconsistent with results of our analyses of GAPDH and other proteins (Figures 4 and 5), it is consistent with relationships seen with some enolase and MDH paralogs (Figure 5).

Bottom Line: As a result, genetic studies of Parabasalia lag behind other organisms.These results are consistent with prior analyses of rDNA and GAPDH sequences and ultrastructural data.These findings, together with the relative ease of Rpb1 isolation, make it an attractive tool for evaluating more extensive relationships within Parabasalia.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Division of Medical Parasitology, New York University School of Medicine, New York, New York, United States of America. sbmalik@dal.ca

ABSTRACT

Background: Parabasalia are single-celled eukaryotes (protists) that are mainly comprised of endosymbionts of termites and wood roaches, intestinal commensals, human or veterinary parasites, and free-living species. Phylogenetic comparisons of parabasalids are typically based upon morphological characters and 18S ribosomal RNA gene sequence data (rDNA), while biochemical or molecular studies of parabasalids are limited to a few axenically cultivable parasites. These previous analyses and other studies based on PCR amplification of duplicated protein-coding genes are unable to fully resolve the evolutionary relationships of parabasalids. As a result, genetic studies of Parabasalia lag behind other organisms.

Principal findings: Comparing parabasalid EF1α, α-tubulin, enolase and MDH protein-coding genes with information from the Trichomonas vaginalis genome reveals difficulty in resolving the history of species or isolates apart from duplicated genes. A conserved single-copy gene encodes the largest subunit of RNA polymerase II (Rpb1) in T. vaginalis and other eukaryotes. Here we directly sequenced Rpb1 degenerate PCR products from 10 parabasalid genera, including several T. vaginalis isolates and avian isolates, and compared these data by phylogenetic analyses. Rpb1 genes from parabasalids, diplomonads, Parabodo, Diplonema and Percolomonas were all intronless, unlike intron-rich homologs in Naegleria, Jakoba and Malawimonas.

Conclusions/significance: The phylogeny of Rpb1 from parasitic and free-living parabasalids, and conserved Rpb1 insertions, support Trichomonadea, Tritrichomonadea, and Hypotrichomonadea as monophyletic groups. These results are consistent with prior analyses of rDNA and GAPDH sequences and ultrastructural data. The Rpb1 phylogenetic tree also resolves species- and isolate-level relationships. These findings, together with the relative ease of Rpb1 isolation, make it an attractive tool for evaluating more extensive relationships within Parabasalia.

Show MeSH
Related in: MedlinePlus