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Large genomic differences between the morphologically indistinguishable diplomonads Spironucleus barkhanus and Spironucleus salmonicida.

Roxström-Lindquist K, Jerlström-Hultqvist J, Jørgensen A, Troell K, Svärd SG, Andersson JO - BMC Genomics (2010)

Bottom Line: Here we address the tempo and mode of such changes within diplomonads, flagellated protists with two nuclei found in oxygen-poor environments.Sequence variations were found between individual S. barkhanus ESTs for many, but not all, protein coding genes.Conversely, no allelic variation was found in a previous genome survey of S. salmonicida.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden.

ABSTRACT

Background: Microbial eukaryotes show large variations in genome structure and content between lineages, indicating extensive flexibility over evolutionary timescales. Here we address the tempo and mode of such changes within diplomonads, flagellated protists with two nuclei found in oxygen-poor environments. Approximately 5,000 expressed sequence tag (EST) sequences were generated from the fish commensal Spironucleus barkhanus and compared to sequences from the morphologically indistinguishable fish parasite Spironucleus salmonicida, and other diplomonads. The ESTs were complemented with sequence variation studies in selected genes and genome size determinations.

Results: Many genes detected in S. barkhanus and S. salmonicida are absent in the human parasite Giardia intestinalis, the most intensively studied diplomonad. For example, these fish diplomonads show an extended metabolic repertoire and are able to incorporate selenocysteine into proteins. The codon usage is altered in S. barkhanus compared to S. salmonicida. Sequence variations were found between individual S. barkhanus ESTs for many, but not all, protein coding genes. Conversely, no allelic variation was found in a previous genome survey of S. salmonicida. This difference was confirmed by sequencing of genomic DNA. Up to five alleles were identified for the cloned S. barkhanus genes, and at least nineteen highly expressed S. barkhanus genes are represented by more than four alleles in the EST dataset. This could be explained by the presence of a non-clonal S. barkhanus population in the culture, by a ploidy above four, or by duplications of parts of the genome. Indeed, genome size estimations using flow cytometry indicated similar haploid genome sizes in S. salmonicida and G. intestinalis (approximately 12 Mb), whereas the S. barkhanus genome is larger (approximately 18 Mb).

Conclusions: This study indicates extensive divergent genome evolution within diplomonads. Genomic traits such as codon usage, frequency of allelic sequence variation, and genome size have changed considerably between S. barkhanus and S. salmonicida. These observations suggest that large genomic differences may accumulate in morphologically indistinguishable eukaryotic microbes.

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UGA serves as a codon for selenocysteine and a polyadenylation signal in addition to termination in S. barkhanus and S. salmonicida. (A) Phylogenetic tree of putative S. barkhanus selenophosphate synthetase with homologs representing the diversity of the protein family. Eukaryotes, Archaea and Bacteria are shown in red, blue and black, respectively. Maximum likelihood bootstrap values and bayesian posterior probabilities are shown above and below branches, respectively. GenBank identification numbers (gi) for each sequence are shown in parentheses. (B) Alignment of a putative S. barkhanus selenoprotein W1 (contig253) with homologs from diplomonads (S. salmonicida - gi:120476942, G. lamblia - gi:253741748, and S. vortens [15]), and a selection of other eukaryotes (Homo sapiens - gi:4506887, Danio rerio - gi:29648542, Chlamydomonas reinhardtii - gi:159471514, Bigelowiella natans - gi:47028259). (C) Sequence logo [54] around the termination codon of 134 S. barkhanus cDNA sequences.
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Figure 1: UGA serves as a codon for selenocysteine and a polyadenylation signal in addition to termination in S. barkhanus and S. salmonicida. (A) Phylogenetic tree of putative S. barkhanus selenophosphate synthetase with homologs representing the diversity of the protein family. Eukaryotes, Archaea and Bacteria are shown in red, blue and black, respectively. Maximum likelihood bootstrap values and bayesian posterior probabilities are shown above and below branches, respectively. GenBank identification numbers (gi) for each sequence are shown in parentheses. (B) Alignment of a putative S. barkhanus selenoprotein W1 (contig253) with homologs from diplomonads (S. salmonicida - gi:120476942, G. lamblia - gi:253741748, and S. vortens [15]), and a selection of other eukaryotes (Homo sapiens - gi:4506887, Danio rerio - gi:29648542, Chlamydomonas reinhardtii - gi:159471514, Bigelowiella natans - gi:47028259). (C) Sequence logo [54] around the termination codon of 134 S. barkhanus cDNA sequences.

Mentions: The third class of genes consists of 58 sequences without matches in the G. intestinalis genome (Additional files 5 and 6). Metabolic functions dominate among the 26 that do have putative annotations, including several peptidases, desulfoferredoxin, fructokinase, cartenoid isomerase and rubrerythrin (Additional file 6), suggesting metabolic adaptation as a selection force for their maintenance in the S. barkhanus genome. Detailed phylogenetic analyses, such as for selenophosphate synthetase (Figure 1A), would be necessary to determine the origin of these genes; they have either been gained in the Spironucleus lineage or been lost in the Giardia lineage. Lateral gene transfer has indeed previously been shown to contribute to adaptation within diplomonads and other eukaryotes [7,14,21,28-30]. In fact, ten of the S. barkhanus sequences have indeed close homologs among proteins identified as recently introduced via lateral gene transfer into the S. salmonicida genome (Additional file 6). Several of the other proteins also likely represent acquisitions in the Spironucleus lineage, some maybe after the divergence between S. barkhanus and S. salmonicida. At any rate, this class contains candidate genes for the understanding of the diversifications of diplomonads, regardless of their origins, as exemplified below.


Large genomic differences between the morphologically indistinguishable diplomonads Spironucleus barkhanus and Spironucleus salmonicida.

Roxström-Lindquist K, Jerlström-Hultqvist J, Jørgensen A, Troell K, Svärd SG, Andersson JO - BMC Genomics (2010)

UGA serves as a codon for selenocysteine and a polyadenylation signal in addition to termination in S. barkhanus and S. salmonicida. (A) Phylogenetic tree of putative S. barkhanus selenophosphate synthetase with homologs representing the diversity of the protein family. Eukaryotes, Archaea and Bacteria are shown in red, blue and black, respectively. Maximum likelihood bootstrap values and bayesian posterior probabilities are shown above and below branches, respectively. GenBank identification numbers (gi) for each sequence are shown in parentheses. (B) Alignment of a putative S. barkhanus selenoprotein W1 (contig253) with homologs from diplomonads (S. salmonicida - gi:120476942, G. lamblia - gi:253741748, and S. vortens [15]), and a selection of other eukaryotes (Homo sapiens - gi:4506887, Danio rerio - gi:29648542, Chlamydomonas reinhardtii - gi:159471514, Bigelowiella natans - gi:47028259). (C) Sequence logo [54] around the termination codon of 134 S. barkhanus cDNA sequences.
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Figure 1: UGA serves as a codon for selenocysteine and a polyadenylation signal in addition to termination in S. barkhanus and S. salmonicida. (A) Phylogenetic tree of putative S. barkhanus selenophosphate synthetase with homologs representing the diversity of the protein family. Eukaryotes, Archaea and Bacteria are shown in red, blue and black, respectively. Maximum likelihood bootstrap values and bayesian posterior probabilities are shown above and below branches, respectively. GenBank identification numbers (gi) for each sequence are shown in parentheses. (B) Alignment of a putative S. barkhanus selenoprotein W1 (contig253) with homologs from diplomonads (S. salmonicida - gi:120476942, G. lamblia - gi:253741748, and S. vortens [15]), and a selection of other eukaryotes (Homo sapiens - gi:4506887, Danio rerio - gi:29648542, Chlamydomonas reinhardtii - gi:159471514, Bigelowiella natans - gi:47028259). (C) Sequence logo [54] around the termination codon of 134 S. barkhanus cDNA sequences.
Mentions: The third class of genes consists of 58 sequences without matches in the G. intestinalis genome (Additional files 5 and 6). Metabolic functions dominate among the 26 that do have putative annotations, including several peptidases, desulfoferredoxin, fructokinase, cartenoid isomerase and rubrerythrin (Additional file 6), suggesting metabolic adaptation as a selection force for their maintenance in the S. barkhanus genome. Detailed phylogenetic analyses, such as for selenophosphate synthetase (Figure 1A), would be necessary to determine the origin of these genes; they have either been gained in the Spironucleus lineage or been lost in the Giardia lineage. Lateral gene transfer has indeed previously been shown to contribute to adaptation within diplomonads and other eukaryotes [7,14,21,28-30]. In fact, ten of the S. barkhanus sequences have indeed close homologs among proteins identified as recently introduced via lateral gene transfer into the S. salmonicida genome (Additional file 6). Several of the other proteins also likely represent acquisitions in the Spironucleus lineage, some maybe after the divergence between S. barkhanus and S. salmonicida. At any rate, this class contains candidate genes for the understanding of the diversifications of diplomonads, regardless of their origins, as exemplified below.

Bottom Line: Here we address the tempo and mode of such changes within diplomonads, flagellated protists with two nuclei found in oxygen-poor environments.Sequence variations were found between individual S. barkhanus ESTs for many, but not all, protein coding genes.Conversely, no allelic variation was found in a previous genome survey of S. salmonicida.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell and Molecular Biology, Uppsala University, Uppsala, Sweden.

ABSTRACT

Background: Microbial eukaryotes show large variations in genome structure and content between lineages, indicating extensive flexibility over evolutionary timescales. Here we address the tempo and mode of such changes within diplomonads, flagellated protists with two nuclei found in oxygen-poor environments. Approximately 5,000 expressed sequence tag (EST) sequences were generated from the fish commensal Spironucleus barkhanus and compared to sequences from the morphologically indistinguishable fish parasite Spironucleus salmonicida, and other diplomonads. The ESTs were complemented with sequence variation studies in selected genes and genome size determinations.

Results: Many genes detected in S. barkhanus and S. salmonicida are absent in the human parasite Giardia intestinalis, the most intensively studied diplomonad. For example, these fish diplomonads show an extended metabolic repertoire and are able to incorporate selenocysteine into proteins. The codon usage is altered in S. barkhanus compared to S. salmonicida. Sequence variations were found between individual S. barkhanus ESTs for many, but not all, protein coding genes. Conversely, no allelic variation was found in a previous genome survey of S. salmonicida. This difference was confirmed by sequencing of genomic DNA. Up to five alleles were identified for the cloned S. barkhanus genes, and at least nineteen highly expressed S. barkhanus genes are represented by more than four alleles in the EST dataset. This could be explained by the presence of a non-clonal S. barkhanus population in the culture, by a ploidy above four, or by duplications of parts of the genome. Indeed, genome size estimations using flow cytometry indicated similar haploid genome sizes in S. salmonicida and G. intestinalis (approximately 12 Mb), whereas the S. barkhanus genome is larger (approximately 18 Mb).

Conclusions: This study indicates extensive divergent genome evolution within diplomonads. Genomic traits such as codon usage, frequency of allelic sequence variation, and genome size have changed considerably between S. barkhanus and S. salmonicida. These observations suggest that large genomic differences may accumulate in morphologically indistinguishable eukaryotic microbes.

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