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    This document discusses the molecular structure of the TaSP gene from Theileria annulata isolates from Sudan. 1) The researchers sequenced and analyzed the polymorphic region of the TaSP gene from T. annulata cell lines and field samples from different areas of Sudan. They found high diversity, with amino acid and length polymorphisms both within and between parasite isolates. 2) Alignment of the sequences showed significant variability, with identities ranging from 41-66% and differences ranging from 16-38%. Variability was observed even within individual animals, suggesting multiple clone infections. 3) The diversity likely allows the parasite to evade host immunity, as TaSP is a highly antigenic protein. However

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    TMP F054

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    This document discusses the molecular structure of the TaSP gene from Theileria annulata isolates from Sudan. 1) The researchers sequenced and analyzed the polymorphic region of the TaSP gene from T. annulata cell lines and field samples from different areas of Sudan. They found high diversity, with amino acid and length polymorphisms both within and between parasite isolates. 2) Alignment of the sequences showed significant variability, with identities ranging from 41-66% and differences ranging from 16-38%. Variability was observed even within individual animals, suggesting multiple clone infections. 3) The diversity likely allows the parasite to evade host immunity, as TaSP is a highly antigenic protein. However

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    This document discusses the molecular structure of the TaSP gene from Theileria annulata isolates from Sudan. 1) The researchers sequenced and analyzed the polymorphic region of the TaSP gene from T. annulata cell lines and field samples from different areas of Sudan. They found high diversity, with amino acid and length polymorphisms both within and between parasite isolates. 2) Alignment of the sequences showed significant variability, with identities ranging from 41-66% and differences ranging from 16-38%. Variability was observed even within individual animals, suggesting multiple clone infections. 3) The diversity likely allows the parasite to evade host immunity, as TaSP is a highly antigenic protein. However

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    50 views3 pages

    TMP F054

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    Frontiers
    This document discusses the molecular structure of the TaSP gene from Theileria annulata isolates from Sudan. 1) The researchers sequenced and analyzed the polymorphic region of the TaSP gene from T. annulata cell lines and field samples from different areas of Sudan. They found high diversity, with amino acid and length polymorphisms both within and between parasite isolates. 2) Alignment of the sequences showed significant variability, with identities ranging from 41-66% and differences ranging from 16-38%. Variability was observed even within individual animals, suggesting multiple clone infections. 3) The diversity likely allows the parasite to evade host immunity, as TaSP is a highly antigenic protein. However

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    of 3

    ANIMAL BIODIVERSITY AND EMERGING DISEASES

    Comparison of the Molecular Structure of the


    TaSP Gene of Theileria annulata from
    Sudanese Isolates
    Awadia M. Ali,a,b Diaeldin Salih,c Mohammed Bakheit,d
    Abdel Rahim M. El Hussein,c Shawgi M. Hassan,a
    Maowia M. Mukhtar,e Jabbar S. Ahmed,b and Ulrike Seitzerb
    a
    Faculty of Veterinary Medicine, University of Khartoum, Khartoum North, Sudan
    b
    Division of Veterinary Infection Biology and Immunology, Research Center Borstel,
    Borstel, Germany
    c
    Central Veterinary Research Laboratories, Al Amarat, Khartoum, Sudan
    d
    National Research Center for Protozoan Diseases, Unit for Diseases Control and
    Genetics, Obihiro University for Agriculture and Veterinary Medicine,
    Inada-cho, Obihiro, Japan
    e
    Institute of Endemic Diseases, University of Khartoum, Khartoum, Sudan

    The polymorphic region of the Theileria annulata surface protein (TaSP) was cloned
    and sequenced from different isolates of cattle and cell lines from different areas of
    Sudan. Amino acid sequence alignment revealed a high diversity showing amino acid
    and length polymorphism, both within and between parasite isolates. The generation
    of TaSP diversity may allow the evasion of host immunity by the parasite since TaSP is
    a highly antigenic parasite protein.

    Key words: alignment; genotype; Theileria annulata; Sudan

    Introduction Materials and Methods

    Theileria annulata, the causative agent of trop- Cell Lines and Field Samples
    ical theileriosis, is a tick-borne protozoan para-
    site. Tropical theileriosis constitutes one of the The parasite isolates and field samples
    obstacles for improved livestock production in from different geographical regions of the
    many countries. In the Sudan, the disease is Sudan used in this study and the number of
    considered as the most important tick-borne clones analyzed per sample are summarized in
    disease.1 No data is available regarding the Table 1.
    genotypes of this Theileria parasite in Sudan.
    This study investigated the molecular struc- Sequence Analysis
    ture of the single copy polymorphic T. annulata
    surface protein (TaSP) gene2 for comparison Genomic DNA was amplified, cloned, and
    of strains isolated from cattle and cell lines in sequenced as described previously.2 Analysis
    Sudan. of the variable region of TaSP was performed
    by comparing the sequences from the various
    isolates (GenBank accession EU032540-
    Address for correspondence: Dr. Ulrike Seitzer, Veterinary Infection EU032577). A total of 38 clones (one to four
    Biology and Immunology, Research Center Borstel, Parkallee 22, 23845
    Borstel, Germany. Voice: +49-(0)4537-188-413; fax +49-(0)4537-188-
    clones of independent PCR reactions) were se-
    627. [email protected] quenced. The predicted amino acid sequences
    Animal Biodiversity and Emerging Diseases: Ann. N.Y. Acad. Sci. 1149: 218–220 (2008).
    doi: 10.1196/annals.1428.025  C 2008 New York Academy of Sciences.

    218
    Ali et al.: Molecular Structure of TaSP Gene of Theileria annulata 219

    TABLE 1. Theileria annulata Isolates and Cell Lines from Different Geographical Locations in the Sudan

    Isolate Origin Type n Clones analyzed

    ATB Atbara (northern Sudan) Cell line, passage 75 1 4


    HAN Hantoub (central Sudan) Cell line, passage 76 1 5
    SHA Shambat (central Sudan) Cell line, passage 10 1 3
    Field samples Khartoum (central Sudan) Field blood samples 14 26

    TABLE 2. Summary of Alignment Results


    Identical Strongly similar Weakly similar Different
    Sample (alignment length) aa (%) aa (%) aa (%) aa (%)

    All sequences, 17 isolates, 38 clones (141) 58 (41.13) 19 (13.48) 11 (7.8) 53 (37.59)


    Field samples, 14 animals, 26 clones (143) 64 (44.76) 17 (11.89) 11 (7.69) 51 (35.66)
    ATB cell line, 4 clones (134) 78 (58.21) 19 (14.18) 8 (5.97) 29 (21.64)
    Field sample, one animal (1109), 4 clones (137) 90 (65.69) 19 (13.87) 6 (4.38) 22 (16.06)

    were aligned using ClustalW available at PBIL similarity 13.87%, weak similarity 4.38%, and
    (Pôle Bio-Informatique Lyonnais) (http:// the difference 16.06%. Alignment of derived
    npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl? amino acid sequences from a single cell line
    page=/NPSA/npsa_clustalw.html).3 (ATB) showed an identity of 78 (58.21%), a
    strong similarity of 19 (14.18%), a weak simi-
    larity of 8 (5.97%), and a difference in amino
    Results acids of 29 (21.64%) over an alignment length
    of 134 amino acids. Alignment results are sum-
    The sequence results showed size and se- marized in Table 2.
    quence polymorphism for the TaSP polymor-
    phic region. Nucleotide sequences were from
    393 to 411 nucleotides, corresponding to de- Discussion
    rived amino acid (aa) sequences of 131 to 137
    residues. Alignment data retrieved from the The results demonstrated a variability of
    ClustalW analysis of all sequences obtained TaSP in Sudanese T. annulata isolates. Interest-
    showed an identity of 41.13% (58 aa residues), ingly, variable sequences were obtained from
    a strong similarity of 13.48% (19 aa residues), a individual animals suggesting multiple clone
    weak similarity of 7.8% (11 aa residues), and infection, which could be a result of the high
    a difference of 37.59% (53 aa residues). There transmission rates that may allow genetic re-
    was high diversity within and between parasite combination of the clones. A similar sequence
    isolates and variable sequences were detected diversity for a gene of T. annulata was previously
    from one animal. No geographical specificity of reported with respect to the major merozoite
    sequence types was observed. Alignment of the antigen gene Tams 1 and also in T. parva for the
    field samples only gave an identity of 44.76%, a polymorphic immunodominant surface pro-
    strong similarity of 11.89%, a weak similarity of tein (PIM), which is closely related to TaSP.2,4,5
    7.69%, and a difference of 35.66%. Using se- No significant association between TaSP se-
    quence data derived from clones obtained from quences and their geographical origins were
    one animal, the identity was 65.69%, strong found. The genetic data of the field isolates
    220 Annals of the New York Academy of Sciences

    indicate a panmictic population structure of Acknowledgments


    the parasite in the Sudan. This result was sup-
    ported by the results obtained from the cell This research was partially supported by
    lines that also showed different alleles in a DAAD (Deutscher Akademischer Austausch
    single culture at passage 75. This result could Dienst), the European Union (ICA4-1999-
    explain the long duration needed to attenu- 30151), Animal Resources Bank (Sudan), and
    ate some T. annulata strains isolated from en- IAEA (International Atomic Energy Agency)
    demic areas, which possibly consist of different through technical project NO.SUD04/
    genotypes. 027 entitled “Control of ticks and tick-borne
    The genetic mechanisms generating the ob- diseases in the Sudan.”
    served polymorphism might be the intragenic
    recombination taking place during the sexual Conflicts of Interest
    phase of the life cycle of the parasite in the The authors declare no conflicts of interest.
    tick vector. Gene conversion, which has been
    identified as a mechanism responsible for the
    reshuffling of important epitopes to create novel References
    alleles, can not be excluded for the TaSP gene,
    although the predominant location of T cell 1. El Hussein, A.M., A.A. Majid & S.M. Hassan. 2004.
    epitopes is within the conserved regions of the The present status of tick-borne diseases in the Sudan.
    molecule.2,6 Gene conversion rather than in- Country report. Arch. Inst. Pasteur Tunis 81: 1–4.
    2. Schnittger, L. et al. 2002. Characterization of a
    tragenic recombination was hypothesized to
    polymorphic Theileria annulata surface protein (TaSP)
    be the main mechanism for the polymor- closely related to PIM of Theileria parva: implications
    phism observed in the T. parva surface protein for use in diagnostic tests and subunit vaccines. Mol.
    (PIM).4 Biochem. Parasitol. 120: 247–256.
    The diversity in antigenic genes is usually the 3. Combet, C. et al. 2000. NPS@: Network Protein Se-
    result of positive selection pressure of the host quence Analysis. TIBS 25: 147–150.
    4. Geysen, D. et al. 2004. An unusual mosaic structure
    immune system, which may also be the case of the PIM gene of Theileria parva and its relationship
    for TaSP, which is a highly antigenic protein.7 to allelic diversity. Mol. Biochem. Parasitol. 133: 163–
    In the TaSP molecule six peptide sequences are 174.
    located in the polymorphic region with binding 5. Katzer, F. et al. 1998. Selection for antigenic diversity
    motifs to bovine major histocompatibility com- of Tams 1, the major merozoite antigen of Theileria
    annulata. Ann. N. Y. Acad. Sci. 849: 96–108.
    plex (MHC) class I molecules.2 Therefore, the
    6. Dormoy, A. et al. 1997. Birth of a new allele in a
    generation of TaSP diversity could also allow sibling: cis or trans gene conversion during meiosis?
    the evasion of host immunity. A previous study Immunogenetics 46: 520–523.
    using merozoite antigen (Tams 1) indicated the 7. Seitzer, U. et al. 2007. From molecule to diagnostic
    selection of parasite genotypes during transmis- tool: Theileria annulata surface protein TaSP. Parasitol.
    sion by ticks, and that selection is mediated via Res. 101(S2): S217–223.
    8. Gubbels, M.J. et al. Study of Theileria annulata popu-
    evasion of the host immune response.8 An in- lation structure during bovine infection and follow-
    vestigation to confirm these possibilities in the ing transmission to ticks. Parasitology 123(Pt 6): 553–
    antigenic gene TaSP is needed in the future. 561.

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