In silico analysis of Ta9 gene polymorphism in an Iranian Theileria annulata schizont-infected cell line S15 vaccine strain and native isolates

Document Type: Original Articles


Department of Parasite Vaccine Research and Production, Razi Vaccine and Serum Research Institute, Karaj, Iran


Bovine theileriosis is a tick-borne disease caused by obligate intracellular parasites related to the genus Theileria. Cellular immune responses protect cattle against pathogens through the activation of immune cells. Nowadays, live, attenuated vaccine of Theileria annulata (T. annulata) is being produced in Iran and is recommended for active cattle immunization. Detection of the immunogenic  antigens and epitopes recognized by CD8+ T Lymphocytes is vital for the development of recombinant and subunit vaccines. Herein, sequences of the genes encoding Ta9, which is an important antigenrecognized by bovine CD8+ T cells specific for T. annulata, in Iranian S15 vaccine strains, several Iranian isolates, as well as reference Ta9 DNA sequences registered in GeneBank were compared through polymerase chain reaction (PCR). The obtained data from DNA sequences were analyzed by using "Nucleotide", "Blast n", "BioEdit" and "IEDB" softwares. The results showed high level of variation in nucleotides and amino acids level. The observed polymorphism in Ta9 gene sequences of Iranian vaccine strains and some isolates from Iran demonstrated that this antigen contains polymorphic sequences and is active along with the specific major histocompatibility complex (MHC) of the host. Polymorphic sequences and specific epitopes of Ta9 gene for CD8+ T cell provides an explanation for incomplete protection observed after inoculation of heterologous parasites in vaccinated cattle. These results have important implications for the application of Ta9 antigen for developing novel subunit vaccines.


Main Subjects

Article Title [French]

Analyse in silico du polymorphisme du gène Ta9 dans une souche iranienne de vaccin S15 infectée par le schizonte Theileria annulata et d’isolats indigènes

Abstract [French]

La theilériose bovine est une infection transmise par des tiques et causée par des parasites obligatoires, intracellulaires appartenant au genre Theileria. La réponse immune cellulaire protège les bovins de ces pathogènes à travers l’activation de cellules immunes spécifiques. Aujourd’hui, le vaccin vivant atténué de Theileria annulata (T. annulata) est produit en Iran et est recommandé pour l’immunisation active des bovins. La détection des antigènes immunogènes et des épitopes reconnus par les lymphocytes T CD8+ est vitale dans le développement de vaccins recombinants ou sous-unitaires. Dans cette étude, les séquences des gènes codant pour Ta9, un antigène majeur reconnu par les cellules T CD8+ bovines et spécifique à l’espèce T. annulata, ont été analysés par réaction en chaine à la polymerase (PCR). Les résultats obtenus pour les souches iraniennes du vaccin S15 et plusieurs isolats iraniens ont été comparés à la séquence ADN de référence du gène Ta9 enregistré dans GeneBank. Les séquences ADN issus de nos analyses ont été étudiées avec les logiciels "Nucleotide", "Blast n", "BioEdit" et "IEDB". Nos résultats montrent un degré important de variation au niveau des séquences nucléotidiques et d’acides aminés. Le polymorphisme observé dans les séquences du gène Ta9 des souches iraniennes du vaccin S15 ainsi que dans plusieurs isolats iraniens montrent que cet antigène contient des séquences fortement polymorphiques et actives au niveau du complexe majeur d’histocompatibilité (CMH) spécifique de l’hôte. Le polymorphisme des séquences et des épitopes spécifiques au gène Ta9 pour les cellules T CD8+ pourrait donc expliquer la protection partielle observée après l’inoculation de parasites hétérologues par les bovins vaccinés. Ces résultats pourraient donc avoir des implications importantes pour le développement de nouveaux vaccins sous-unitaires à partir de l’antigène Ta9.

Keywords [French]

  • Antigène
  • Iran
  • Polymorphisme
  • Ta9
  • Theileria annulata
  • Vaccin
Ahmed, J.S., Hartwig, H., Schein, E., 1999. Generation of Theileria annulata-specific cytotoxic T lymphocytes coincides with the control of tropical theileriosis. Parasitol Res 85, 870-872.

Campbell, J.D., Russell, G.C., Nelson, R.E., Spooner, R.L., Glass, E.J., 1997. Theileria annulata "superantigen" activity--TCB usage by responding bovine CD4+ cells from uninfected donors. Biochem Soc Trans 25, 277S.

Darghouth, M.A., Ben Miled, L., Bouattour, A., Melrose, T.R., Brown, C.G., Kilani, M., 1996. A preliminary study on the attenuation of Tunisian schizont-infected cell lines of Theileria annulata. Parasitol Res 82, 647-655.

Emery, D.L., Eugui, E.M., Nelson, R.T., Tenywa, T., 1981. Cell-mediated immune responses to Theileria parva (East Coast fever) during immunization and lethal infections in cattle. Immunology 43, 323-336.

Goddeeris, B.M., Morrison, W.I., Toye, P.G., Bishop, R., 1990. Strain specificity of bovine Theileria parva-specific cytotoxic T cells is determined by the phenotype of the restricting class I MHC. Immunology 69, 38-44.

Graham, S.P., Pelle, R., Yamage, M., Mwangi, D.M., Honda, Y., Mwakubambanya, R.S., de Villiers, E.P., Abuya, E., Awino, E., Gachanja, J., Mbwika, F., Muthiani, A.M., Muriuki, C., Nyanjui, J.K., Onono, F.O., Osaso, J., Riitho, V., Saya, R.M., Ellis, S.A., McKeever, D.J., MacHugh, N.D., Gilbert, S.C., Audonnet, J.C., Morrison, W.I., van der Bruggen, P., Taracha, E.L., 2008. Characterization of the fine specificity of bovine CD8 T-cell responses to defined antigens from the protozoan parasite Theileria parva. Infect Immun 76, 685-694.

Habibi, G., 2012. Phylogenetic Analysis of Theileria annulata Infected Cell Line S15 Iran Vaccine Strain. Iran J Parasitol 7, 73-81.

Hansen, A.M., Rasmussen, M., Svitek, N., Harndahl, M., Golde, W.T., Barlow, J., Nene, V., Buus, S., Nielsen, M., 2014. Characterization of binding specificities of bovine leucocyte class I molecules: impacts for rational epitope discovery. Immunogenetics 66, 705-718.

Hayashida, K., Abe, T., Weir, W., Nakao, R., Ito, K., Kajino, K., Suzuki, Y., Jongejan, F., Geysen, D., Sugimoto, C., 2013. Whole-genome sequencing of Theileria parva strains provides insight into parasite migration and diversification in the African continent. DNA Res 20, 209-220.

Jappe, U., 2000. Superantigens and their association with dermatological inflammatory diseases: facts and hypotheses. Acta Derm Venereol 80, 321-328.

Machugh, N.D., Burrells, A.C., Morrison, W.I., 2008. Demonstration of strain-specific CD8 T cell responses to Theileria annulata. Parasite Immunol 30, 385-393.

MacHugh, N.D., Weir, W., Burrells, A., Lizundia, R., Graham, S.P., Taracha, E.L., Shiels, B.R., Langsley, G., Morrison, W.I., 2011. Extensive polymorphism and evidence of immune selection in a highly dominant antigen recognized by bovine CD8 T cells specific for Theileria annulata. Infect Immun 79, 2059-2069.

McKeever, D.J., Taracha, E.L., Innes, E.L., MacHugh, N.D., Awino, E., Goddeeris, B.M., Morrison, W.I., 1994. Adoptive transfer of immunity to Theileria parva in the CD8+ fraction of responding efferent lymph. Proc Natl Acad Sci U S A 91, 1959-1963.

Morrison, W.I., 1996. Influence of host and parasite genotypes on immunological control of Theileria parasites. Parasitology 112 Suppl, S53-66.

Morrison, W.I., Connelley, T., Hemmink, J.D., MacHugh, N.D., 2015. Understanding the basis of parasite strain-restricted immunity to Theileria parva. Annu Rev Anim Biosci 3, 397-418.

Morrison, W.I., Goddeeris, B.M., Teale, A.J., Groocock, C.M., Kemp, S.J., Stagg, D.A., 1987. Cytotoxic T-cells elicited in cattle challenged with Theileria parva (Muguga): evidence for restriction by class I MHC determinants and parasite strain specificity. Parasite Immunol 9, 563-578.

Morrot, A., Zavala, F., 2004. Effector and memory CD8+ T cells as seen in immunity to malaria. Immunol Rev 201, 291-303.

Pipano, E., 1989. Vaccination against Theileria annulata theileriosis, CRC Press, Inc., Boca Raton, FL.

Preston, P.M., Brown, C.G., Spooner, R.L., 1983. Cell-mediated cytotoxicity in Theileria annulata infection of cattle with evidence for BoLA restriction. Clin Exp Immunol 53, 88-100.

Radley, D.E., Brown, C.G.D., Burridge, M.J., Cunningham, M.P., Kirimi, I.M., Purnell, R.E., Young, A.S., 1975. East coast fever: 1. Chemoprophylactic immunization of cattle against Theileria parva (Muguga) and five theilerial strains. Veterinary Parasitology 1, 35-41.

Sambrook, J., Fritsch, E.F., Maniatis, T., 1989. Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor; NY, B.16.

Spooner, R.L., Innes, E.A., Glass, E.J., Brown, C.G., 1989. Theileria annulata and T. parva infect and transform different bovine mononuclear cells. Immunology 66, 284-288.

Sutherland, I.A., Shiels, B.R., Jackson, L., Brown, D.J., Brown, C.G., Preston, P.M., 1996. Theileria annulata: altered gene expression and clonal selection during continuous in vitro culture. Exp Parasitol 83, 125-133.

Taracha, E.L., Goddeeris, B.M., Teale, A.J., Kemp, S.J., Morrison, W.I., 1995. Parasite strain specificity of bovine cytotoxic T cell responses to Theileria parva is determined primarily by immunodominance. J Immunol 155, 4854-4860.

Weir, W., Ben-Miled, L., Karagenc, T., Katzer, F., Darghouth, M., Shiels, B., Tait, A., 2007. Genetic exchange and sub-structuring in Theileria annulata populations. Mol Biochem Parasitol 154, 170-180.