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Draft genome sequencing of giardia intestinalis assemblage B isolate GS: is human giardiasis caused by two different species?

Franzén O, Jerlström-Hultqvist J, Castro E, Sherwood E, Ankarklev J, Reiner DS, Palm D, Andersson JO, Andersson B, Svärd SG - PLoS Pathog. (2009)

Bottom Line: We here performed 454 sequencing to 16x coverage of the assemblage B isolate GS, the only Giardia isolate successfully used to experimentally infect animals and humans.The tetraploid GS genome shows higher levels of overall allelic sequence polymorphism (0.5 versus <0.01% in WB).The genomic differences between WB and GS may explain some of the observed biological and clinical differences between the two isolates, and it suggests that assemblage A and B Giardia can be two different species.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
Giardia intestinalis is a major cause of diarrheal disease worldwide and two major Giardia genotypes, assemblages A and B, infect humans. The genome of assemblage A parasite WB was recently sequenced, and the structurally compact 11.7 Mbp genome contains simplified basic cellular machineries and metabolism. We here performed 454 sequencing to 16x coverage of the assemblage B isolate GS, the only Giardia isolate successfully used to experimentally infect animals and humans. The two genomes show 77% nucleotide and 78% amino-acid identity in protein coding regions. Comparative analysis identified 28 unique GS and 3 unique WB protein coding genes, and the variable surface protein (VSP) repertoires of the two isolates are completely different. The promoters of several enzymes involved in the synthesis of the cyst-wall lack binding sites for encystation-specific transcription factors in GS. Several synteny-breaks were detected and verified. The tetraploid GS genome shows higher levels of overall allelic sequence polymorphism (0.5 versus <0.01% in WB). The genomic differences between WB and GS may explain some of the observed biological and clinical differences between the two isolates, and it suggests that assemblage A and B Giardia can be two different species.

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Comparison of encystation-specific promoters.The promoters of the cyst-wall proteins-1 and 2 (CWP-1 and CWP-2), the encystation-specific transcription factor (gMyb2), the key enzymes involved in the synthesis of cyst wall sugars, glucosamine-6 phosphate isomerase (G6PI-B) and UDP-glucosamine-4 epimerase (UG4E) were aligned using the program CLUSTALW. Poly A rich initiator regions and gMyb2 binding-sites are underlined. Note that the G6PI-B and UG4E promoters are missing the gMyb2 binding-sites, whereas they are found in the corresponding positions in the promoters of CWP-1 and –2 and gMyb2.
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ppat-1000560-g004: Comparison of encystation-specific promoters.The promoters of the cyst-wall proteins-1 and 2 (CWP-1 and CWP-2), the encystation-specific transcription factor (gMyb2), the key enzymes involved in the synthesis of cyst wall sugars, glucosamine-6 phosphate isomerase (G6PI-B) and UDP-glucosamine-4 epimerase (UG4E) were aligned using the program CLUSTALW. Poly A rich initiator regions and gMyb2 binding-sites are underlined. Note that the G6PI-B and UG4E promoters are missing the gMyb2 binding-sites, whereas they are found in the corresponding positions in the promoters of CWP-1 and –2 and gMyb2.

Mentions: Promoters in Giardia are short (around 50 bp) and the main feature is an initiator-like AT-rich sequence around the ATG start codon, which is enough to drive transcription [32]. AT-rich stretches around the ATG start codon could be found in most GS genes (data not shown), but apart from these, there is very little intergenic sequence conservation, consistent with earlier observations of a few GS genes. Encystation-specific promoters in WB are also short, with 65 bp found to be sufficient for a developmentally regulated promoter [33]. An alignment of the promoters (−100 to +3) from the three major cyst-wall proteins CWP 1–3 from WB with the orthologs from GS showed a high degree of conservation in the 65 bp directly upstream of the start codon (see alignment of CWP 1 and 2, Fig. 4). The transcription factor Myb2 has been shown to bind to the CWP promoters and its own promoter [34]. The GS Myb2 protein is well conserved (77% amino acid identity) and so is the 65 bp directly upstream of the ATG start-codon, including the Myb2 binding site (Fig. 4). This suggests that the cyst-wall proteins and Myb2 protein are regulated in the same way during encystation in the two isolates. The key-regulatory enzyme in WB, glucosamine-6 phosphate isomerase, has a promoter that is similar to the cyst-wall promoters [33]. However, the promoter of this enzyme in GS is not similar to the promoters of the cyst wall proteins (Fig. 4) and, most importantly, it lacks a typical Myb2 binding sequence. The same was found to be true for the last enzyme in the pathway, UDP-N-acetylglucosamine 4′ epimerase [35] (Fig. 4). The GS isolate is known for its poor encysting ability in vitro [36] and may suggest that the regulation of cyst-wall sugar synthesis during early encystation is different in GS.


Draft genome sequencing of giardia intestinalis assemblage B isolate GS: is human giardiasis caused by two different species?

Franzén O, Jerlström-Hultqvist J, Castro E, Sherwood E, Ankarklev J, Reiner DS, Palm D, Andersson JO, Andersson B, Svärd SG - PLoS Pathog. (2009)

Comparison of encystation-specific promoters.The promoters of the cyst-wall proteins-1 and 2 (CWP-1 and CWP-2), the encystation-specific transcription factor (gMyb2), the key enzymes involved in the synthesis of cyst wall sugars, glucosamine-6 phosphate isomerase (G6PI-B) and UDP-glucosamine-4 epimerase (UG4E) were aligned using the program CLUSTALW. Poly A rich initiator regions and gMyb2 binding-sites are underlined. Note that the G6PI-B and UG4E promoters are missing the gMyb2 binding-sites, whereas they are found in the corresponding positions in the promoters of CWP-1 and –2 and gMyb2.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2723961&req=5

ppat-1000560-g004: Comparison of encystation-specific promoters.The promoters of the cyst-wall proteins-1 and 2 (CWP-1 and CWP-2), the encystation-specific transcription factor (gMyb2), the key enzymes involved in the synthesis of cyst wall sugars, glucosamine-6 phosphate isomerase (G6PI-B) and UDP-glucosamine-4 epimerase (UG4E) were aligned using the program CLUSTALW. Poly A rich initiator regions and gMyb2 binding-sites are underlined. Note that the G6PI-B and UG4E promoters are missing the gMyb2 binding-sites, whereas they are found in the corresponding positions in the promoters of CWP-1 and –2 and gMyb2.
Mentions: Promoters in Giardia are short (around 50 bp) and the main feature is an initiator-like AT-rich sequence around the ATG start codon, which is enough to drive transcription [32]. AT-rich stretches around the ATG start codon could be found in most GS genes (data not shown), but apart from these, there is very little intergenic sequence conservation, consistent with earlier observations of a few GS genes. Encystation-specific promoters in WB are also short, with 65 bp found to be sufficient for a developmentally regulated promoter [33]. An alignment of the promoters (−100 to +3) from the three major cyst-wall proteins CWP 1–3 from WB with the orthologs from GS showed a high degree of conservation in the 65 bp directly upstream of the start codon (see alignment of CWP 1 and 2, Fig. 4). The transcription factor Myb2 has been shown to bind to the CWP promoters and its own promoter [34]. The GS Myb2 protein is well conserved (77% amino acid identity) and so is the 65 bp directly upstream of the ATG start-codon, including the Myb2 binding site (Fig. 4). This suggests that the cyst-wall proteins and Myb2 protein are regulated in the same way during encystation in the two isolates. The key-regulatory enzyme in WB, glucosamine-6 phosphate isomerase, has a promoter that is similar to the cyst-wall promoters [33]. However, the promoter of this enzyme in GS is not similar to the promoters of the cyst wall proteins (Fig. 4) and, most importantly, it lacks a typical Myb2 binding sequence. The same was found to be true for the last enzyme in the pathway, UDP-N-acetylglucosamine 4′ epimerase [35] (Fig. 4). The GS isolate is known for its poor encysting ability in vitro [36] and may suggest that the regulation of cyst-wall sugar synthesis during early encystation is different in GS.

Bottom Line: We here performed 454 sequencing to 16x coverage of the assemblage B isolate GS, the only Giardia isolate successfully used to experimentally infect animals and humans.The tetraploid GS genome shows higher levels of overall allelic sequence polymorphism (0.5 versus <0.01% in WB).The genomic differences between WB and GS may explain some of the observed biological and clinical differences between the two isolates, and it suggests that assemblage A and B Giardia can be two different species.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Molecular Biology, Karolinska Institutet, Stockholm, Sweden.

ABSTRACT
Giardia intestinalis is a major cause of diarrheal disease worldwide and two major Giardia genotypes, assemblages A and B, infect humans. The genome of assemblage A parasite WB was recently sequenced, and the structurally compact 11.7 Mbp genome contains simplified basic cellular machineries and metabolism. We here performed 454 sequencing to 16x coverage of the assemblage B isolate GS, the only Giardia isolate successfully used to experimentally infect animals and humans. The two genomes show 77% nucleotide and 78% amino-acid identity in protein coding regions. Comparative analysis identified 28 unique GS and 3 unique WB protein coding genes, and the variable surface protein (VSP) repertoires of the two isolates are completely different. The promoters of several enzymes involved in the synthesis of the cyst-wall lack binding sites for encystation-specific transcription factors in GS. Several synteny-breaks were detected and verified. The tetraploid GS genome shows higher levels of overall allelic sequence polymorphism (0.5 versus <0.01% in WB). The genomic differences between WB and GS may explain some of the observed biological and clinical differences between the two isolates, and it suggests that assemblage A and B Giardia can be two different species.

Show MeSH
Related in: MedlinePlus