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Genomic variation within Giardia intestinalis assemblage A isolates
Uppsala University, Disciplinary Domain of Science and Technology. (Prof Staffan Svärd Laboratory)
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Background: The diarrhea-causing protozoan Giardia intestinalis makes up a species complex of eight different assemblages (A-H), where assemblage A and B infect humans. We have performed whole genome sequencing of two sub-assemblage AII isolates, recently axenized from symptomatic patients, to study the genetic diversity within assemblage A and to identify new assemblage A-specific genotyping targets.

Results: Several biological differences between the assemblage A isolates were identified, including a difference in growth medium preference. The two AII isolates were of different sub-assemblage types (AII-1 (AS98) and AII-2 (AS175)) and showed size differences in the smallest chromosomes. The amount of genetic diversity was characterized in relation to the genome of an assemblage AI isolate (WB). Our analyses indicate that the divergence between AI and AII is approximately 1%, represented by ~100,000 single nucleotide polymorphisms (SNP). Moreover, SNPs are homogeneously distributed over the chromosomes with an enrichment in regions containing surface antigens and non-coding sequences. The level of allelic sequence heterozygosity (ASH) in the two AII isolates were found to be 0.25-0.35%, which is 25-30-fold higher than in the WB isolate. 37 proteinencoding genes, not found in the WB genome, were identified in the two AII genomes. The large gene families of variant-specific surface proteins (VSPs) and high cysteine membrane proteins (HCMPs) showed isolatespecific divergences of the gene repertoires. Certain genes, often in small gene families with 2 to 7 members, showed high sequence diversity between the assemblage A isolates and they could have important roles in hostparasite interactions. A subset of the variable genes was used to develop new genotyping methods for assemblage A isolates.

Conclusions: Our results show that there is a significant genomic variation in assemblage A isolates, in terms of chromosome size, gene content, surface protein repertoire and gene polymorphisms. This identified putative virulence genes and generated a new assemblage A-specific genotyping approach.

Keyword [en]
Giardia, comparative genomics, assemblage A, ASH, MLG
National Category
Medical and Health Sciences Natural Sciences
Research subject
Biology with specialization in Microbiology; Molecular Medicine; Infectious Diseases
URN: urn:nbn:se:uu:diva-167052OAI: oai:DiVA.org:uu-167052DiVA: diva2:480627
Available from: 2012-01-19 Created: 2012-01-19 Last updated: 2012-02-15
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.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 893
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
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)
Available from: 2012-02-02 Created: 2012-01-19 Last updated: 2012-02-15Bibliographically approved

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