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Genome analysis and comparative genomics of a Giardia intestinalis assemblage E isolate.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Evolution, Genomics and Systematics.
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2010 (English)In: BMC Genomics, ISSN 1471-2164, E-ISSN 1471-2164, Vol. 11, 543- p.Article in journal (Refereed) Published
Abstract [en]

Background

Giardia intestinalis is a protozoan parasite that causes diarrhea in a wide range of mammalian species. To further understand the genetic diversity between the Giardia intestinalis species, we have performed genome sequencing and analysis of a wild-type Giardia intestinalis sample from the assemblage E group, isolated from a pig.

Results

We identified 5012 protein coding genes, the majority of which are conserved compared to the previously sequenced genomes of the WB and GS strains in terms of microsynteny and sequence identity. Despite this, there is an unexpectedly large number of chromosomal rearrangements and several smaller structural changes that are present in all chromosomes. Novel members of the VSP, NEK Kinase and HCMP gene families were identified, which may reveal possible mechanisms for host specificity and new avenues for antigenic variation. We used comparative genomics of the three diverse Giardia intestinalis isolates P15, GS and WB to define a core proteome for this species complex and to identify lineage-specific genes. Extensive analyses of polymorphisms in the core proteome of Giardia revealed differential rates of divergence among cellular processes.

Conclusions

Our results indicate that despite a well conserved core of genes there is significant genome variation between Giardia isolates, both in terms of gene content, gene polymorphisms, structural chromosomal variations and surface molecule repertoires. This study improves the annotation of the Giardia genomes and enables the identification of functionally important variation.

Place, publisher, year, edition, pages
2010. Vol. 11, 543- p.
National Category
Biological Sciences Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-133827DOI: 10.1186/1471-2164-11-543ISI: 000283123400001PubMedID: 20929575OAI: oai:DiVA.org:uu-133827DiVA: diva2:371256
Available from: 2010-11-19 Created: 2010-11-16 Last updated: 2017-12-12Bibliographically approved
In thesis
1. Inter and Intra-Assemblage Characterizations of Giardia intestinalis: from clinic to genome
Open this publication in new window or tab >>Inter and Intra-Assemblage Characterizations of Giardia intestinalis: from clinic to genome
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The protozoan parasite Giardia intestinalis (syn. G. lamblia, G. duodenalis) is one of the most common causes of diarrheal disease throughout the world, where an estimated 500 million people are infected annually. Despite efforts in trying to elucidate factors associated with virulence in G. intestinalis little is currently known. The disease outcome is highly variable in Giardia infected individuals, ranging from asymptomatic carriers to severe disease. The reasons behind the differences in disease outcome are vaguely understood and studies trying to link infectivity to different Giardia assemblages or sub-assemblages have rendered conflicting results. Prior to this study, little was known about the prevalence and genetic diversity of different G. intestinalis assemblages across the world.

In this thesis, molecular characterization of clinical G. intestinalis samples from Eastern Africa and Central America, has been performed, enabling a better understanding of the prevalence of different Giardia genotypes in endemic areas (Papers I and II). A correlation between Giardia colonization and the presence of Helicobacter pylori in the human host was established. We found that the currently available genotyping tools provide low resolution when used to characterize assemblage A Giardia. Also, genotyping of assemblage B isolates at these loci is troublesome due to the polymorphic substitutions frequently found in the sequencing chromatograms. This ambiguity was investigated by using micromanipulation to isolate single assemblage B Giardia cells (Paper III). Both cultured trophozoites and cysts from giardiasis patients were analyzed. The data showed that allelic sequence heterozygosity (ASH) does occur at the single cell level, but also that multiple sub-assemblage infections appear to be common in human giardiasis patients.

Furthermore, genome-wide sequencing followed by comparative genomics was performed in order to better characterize differences between and within different Giardia assemblages. The genome of a non-human infecting, assemblage E isolate (Paper IV) was sequenced.  The genomes of two freshly isolated human infecting assemblage AII isolates were also sequenced (Paper V). Subsequent, comparative analyses were performed and included the genomes of two human infecting isolates, WB (AI) and GS/M (B). Several important differences were found between assemblages A, B and E, but also within assemblage A; including unique gene repertoires for each isolate, observed differences in the variable gene families and an overall difference in ASH between the different isolates. Also, a new multi-locus genotyping (MLG) strategy for genotyping of assemblage A Giardia has been established and evaluated on clinical samples from human giardiasis patients.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2012. 85 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 893
Keyword
Giardia, protozoa, parasite infection, diarrhea, genome sequencing, comparative genomics, genotyping, ASH, MLG
National Category
Natural Sciences Medical and Health Sciences
Research subject
Infectious Diseases; Molecular Medicine; Microbiology
Identifiers
urn:nbn:se:uu:diva-167063 (URN)978-91-554-8259-6 (ISBN)
Public defence
2012-02-23, B42, BMC, Husargatan 3, Uppsala, 10:15 (English)
Opponent
Supervisors
Available from: 2012-02-02 Created: 2012-01-19 Last updated: 2012-02-15Bibliographically approved
2. Hidden Diversity Revealed: Genomic, Transcriptomic and Functional Studies of Diplomonads
Open this publication in new window or tab >>Hidden Diversity Revealed: Genomic, Transcriptomic and Functional Studies of Diplomonads
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The diplomonads are a diverse group of eukaryotic microbes found in oxygen limited environments such as the intestine of animals were they may cause severe disease. Among them, the prominent human parasite Giardia intestinalis non-invasively colonizes the small intestine of humans and animals where it induces the gastrointestinal disease giardiasis. Two of the eight genetic groups of G. intestinalis, assemblage A and B, are known to infect humans and have zoonotic potential. At the start of project, genome scale data from assemblage B-H was either sparse or entirely missing.

In this thesis, genome sequencing was performed on the assemblage B isolate GS (Paper I) and the P15 isolate (Paper III) of the hoofed-animals specific assemblage E to investigate the underlying components of phenotypic diversity in Giardia. Comparisons to assemblage A isolate WB revealed large genomic differences; entirely different repertoires of surface antigens, genome rearrangements and isolate specific coding sequences of potential bacterial origin. We established that genomic differences are also manifested at the transcriptome level (Paper VIII). In a follow up analysis (Paper IV) we concluded that the Giardia assemblages are largely reproductively isolated. The large genomic differences observed between Giardia isolates can explain the phenotypic diversity of giardiasis.

The adaptation of diplomonads was further studied in Spironucleus barkhanus (Paper II), a fish commensal of grayling, that is closely related to the fish pathogen Spironucleus salmonicida, causative agent of systemic spironucleosis in salmonid fish. We identified substantial genomic differences in the form of divergent genome size, primary sequence divergence and evidence of allelic sequence heterozygosity, a feature not seen in S. salmonicida.

We devised a transfection system for S. salmonicida (Paper VI) and applied it to the study of the mitochondrial remnant organelle (Paper VII). Our analyses showed that S. salmonicida harbor a hydrogenosome, an organelle with more metabolic capabilities than the mitosome of Giardia. Phylogenetic reconstructions of key hydrogenosomal enzymes showed an ancient origin, indicating a common origin to the hydrogenosome in parabasilids and diplomonads.

In conclusion, the thesis has provided important insights into the adaptation of diplomonads in the present and the distant past, revealing hidden diversity.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2012. 104 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 990
Keyword
Giardia intestinalis, Spironucleus salmonicida, Spironucleus barkhanus, intestinal parasite, hydrogenosome, mitosome, lateral gene transfer, horizontal gene transfer, diplomonad, metamonad, sexual recombination, transfection, protein complex purification
National Category
Microbiology Evolutionary Biology Infectious Medicine
Research subject
Biology with specialization in Evolutionary Organismal Biology; Biology with specialization in Microbiology; Biology with specialization in Molecular Biology; Biology with specialization in Molecular Evolution
Identifiers
urn:nbn:se:uu:diva-182831 (URN)978-91-554-8520-7 (ISBN)
Public defence
2012-12-14, B22, Biomedicinskt centrum (BMC), Husargatan 3, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2012-11-22 Created: 2012-10-16 Last updated: 2013-02-11Bibliographically approved

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