Iranian Journal of Parasitology 2015. 10(4):590-598.

Evaluation of Immunogenicity of Cocktail DNA Vaccine Contain¬ing Plasmids Encoding Complete GRA5, SAG1, and ROP2 Antigens of Toxoplasma gondii in BALB/C Mice


Background: Severe and fatal complications of toxoplasmosis urge development of effective vaccines against the disease. The current study was performed to evalu­ate cocktail DNA vaccine containing plasmids encoding GRA5, SAG1, and ROP2 genes of Toxoplasma gondii in BALB/c mice in Tarbiat Modares University in 2012.

Methods: The plasmids containing complete GRA5, SAG1, and ROP2 genes were mass extracted and then the recombinant plasmids were administered via intramuscu­lar injections according to immunized mice three times with three-week intervals. Then splenocytes were cultured, and proliferation as well as cytokine as­says were carried out. The other mice in each group were inoculated by the parasite and mortality of the mice was evaluated on a daily basis.

Results: The results of cytokine assay for INF-γ were higher in the mice that re­ceived the cocktail DNA containing recombinant plasmids. Evaluation of prolifera­tion of splenocytes using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazo­lium bromide) assay indicated induction of cellular response. Measurement of total IgG and the isotypes of IgG1 and IgG2a showed that the cocktail DNA stimulated IgG and IgG2a production in comparison with the control groups (P<0.05). Furthermore, the survival rate of mice in the groups that received the cocktail DNA was significantly higher than that in the control groups (P<0.05).

Conclusion: Administration of the cocktail DNA vaccine led to production of higher levels of IFN-γ, confirmed by secretion of IgG2a, and the immune response was shifted toward Th1. Thus, the cocktail DNA containing the recombinant plas­mids can be an appropriate candidate for immunization against toxoplasmosis.


Toxoplasma gondii; Cocktail DNA Vac-cine; pcGRA5; pcSAG1; pcROP2

Full Text:



Araujo FG. Immunization against Toxoplasma gondii. Parasitol Today. 1994; 10:358–360.

Couvreur G, Sadak A, Fortier B, Dubremetz JF. Surface antigens of Toxoplasma gondii. Parasitol-ogy. 1988; 97:1–10.

Capron A, Dessaint JP. Vaccination against par-asitic diseases, some alternative concepts for the definition of protective antigens. Ann Inst Pas-teur Immunol. 1988;139:109-117.

Cesbron MF, Capran A. Excreted-secreted anti-gens of Toxoplasma gondii: their origin and role in the host parasite interaction. Res Immunol. 1993;144:41-47.

Cesbron-Delauw MF. Dense granule organelles of Toxoplasma gondii: their role in the host parasite relationship. Parasitol Today. 1994; 10(8):293-296.

Saavedra R, Becerril MA, Dubeaux C, Lippens R, Vos MD, Herion P, Bollen A. Epitopes rec-ognized by human T lymphocytes in the RoP2 protein antigen of Toxoplasma gondii. Infect Im-mun. 1996; 64:3858–3862.

Sadak A, Taghy Z, Fortier B, Dubremetz JF. Characterization of a family of rhoptry proteins of Toxoplasma gondii. Mol Biochem Parasitol. 1998; 29: 203-211.

Velge-Roussel F, Marcelo P, Lepage AC, Buzoni-Gatel D, Bout D. Intranasal immuniza-tion with Toxoplasma gondii SAG1 induces pro-tective cells into NALT and GALT compart-ments. Infec Immun. 2000; 68(2):969-972.

Nielsen HV, Lauemoller SL, Christiansen L, Buus S, Fomsgaard A, Petersen E. Complete protection against lethal Toxoplasma gondii infec-tion in miceimmunized with a plasmid encoding the SAG1 gene. Infect Immun. 1999; 67(12):6358-63.

Liu KY, Zhang DB, Wei QK, Li J, Li GP, Yu JZ. Biological role of surface Toxoplasma gondii antigen in development of vaccine. World J Gastroenterol. 2006; 2363:8-12.

Solhjoo K, Ghaffari far F, Dalimi A, Sharifi Z. Enhancement of antibody immune response to a Toxoplasma gondii SAG1-encoded DNA vac-cine by formulation with aluminium phosphate. J Med Sci. 2007; 7(3):361-367.

Hoseinian Khosroshahi K, Ghaffarifar F, D’Souza S, Sharifi Z, Dalimi A. Evaluation of

the immune response induced by DNA vaccine cocktail expressing complete SAG1 and ROP2 genes against Toxoplasma gondii. Vaccine. 2011; 29:778-783.

Holec-Gasior L, Kur J. Toxoplasma gondii: Re-combinant GRA5 antigen for detection of im-munoglobulin G antibodies using enzyme-linked immunosorbent assay. Exp Parasitol. 2010; 124(3): 272-8.

Igarashi M, Kano F, Tamekuni K, Kawasaki PM, Navarro IT, Vidotto O, et al. Toxoplasma gondii: cloning, sequencing, expression, and anti-genic characterization of ROP2, GRA5 and GRA7. Genet Mol Res. 2008; 7(2): 305-13.

Hosseinian Khosroshahi K, Ghaffarifar F, Shar-ifi Z, Dalimi A. Expression of complete rhoptry protein 2 (ROP2) gene of Toxoplasma gondii in eukaryotic cell. Afr J Biotechnol. 2008; 7(24):4432–6.

Naserifar R , Ghaffarifar F, Dalimi A, Sharifi Z. Cloning of Toxoplasma gondii Granular Antigen 5(GRA5) in Expression Eukaryotic Plasmid pcDNA3 and its Expression on CHO Cell.2011. J Ilam Univ Med Sci. 19(3):1-12.

Solhjoo K, Ghaffari far, F, Dalimi A, Sharifi Z. Cloning and sequencing of Toxoplasma gondii ma-jor surface antigen (SAG1) gene. Arch Razi Inst. 2006; 61(2):73-79.

Holec-Gasior L, Kur J. Toxoplasma gondii: Re-combinant GRA5 antigen for detection of im-munoglobulin G antibodies using enzyme-linked immunosorbent assay. Exp Parasitol. 2010; 124(3): 272-8.

Igarashi M, Kano F, Tamekuni K, Machado RZ, Navarro IT, Vidotto O, et al. Toxoplasma gondii: Evaluation of an intranasal vaccine using recombinant proteins against brain cyst formation in BALB/c mice. Exp Parasitol. 2008;118(3):386-92.

Zenner L, Estaquier J, Darcy F, Maes P, Capron A, Cesbron-Delauw MF. Protective immunity in the rat model of congenital toxoplasmosis and the potential of excreted-secreted antigens as vaccine components. Parasite Immunol. 1999; 21(5):261-72.


  • There are currently no refbacks.

Creative Commons Attribution-NonCommercial 3.0

This work is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License which allows users to read, copy, distribute and make derivative works for non-commercial purposes from the material, as long as the author of the original work is cited properly.