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CLONING AND EXPRESSION OF TOXOPLASMA GONDII DENSE GRANULE ANTIGEN 2 (GRA2) GENE BY PICHIA PASTORIS

Department of Parasitology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
The Southeast Asian journal of tropical medicine and public health (Impact Factor: 0.61). 01/2012;

ABSTRACT Detection of Toxoplasma gondii infection is essential in pregnant women and immunosuppressed patients. Numerous studies have shown that the recombi-nant production of several Toxoplasma antigens, including dense granule antigens (GRAs) has high potential as diagnostic reagents. In the present study, we pro-duced GRA2 using Pichia pastoris system. RNA of T. gondii RH strain tachyzoite was used as a template to produce cDNA clones of full-length GRA2 via reverse transcriptase PCR. Amplicons were inserted into pPICZa A and the recombinant plasmid transformed into P. pastoris, X-33 strain. The expressed recombinant pro-tein was identified by SDS-PAGE and Western blotting. A recombinant protein of ~28 kDa was produced, which could be detected by toxoplasmosis positive human sera indicating that the recombinant protein retained its antigenicity. The present study indicates that P. pastoris-expressed GRA2 should be useful for detection of Toxoplasma infection.

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    ABSTRACT: Toxoplasma gondii is a parasitic protozoan which is the cause of disease toxoplasmosis. Although human toxoplasmosis in healthy adults is usually asymptomatic, a serious disease can occur in the case of a congenital infection and immunocompromised individuals. Furthermore, despite the exact recognition of its etiology, it still presents a diagnostic problem. Diagnosis of toxoplasmosis is mainly based on the results of serological tests detecting anti-T. gondii specific antibodies in the patient's serum sample. The specificities and sensitivities of serology tests depend mostly on the diagnostic antigen(s) used. Most of commercial serological kits currently available are based on Toxoplasma lysate antigens (TLAs). In recent years, many studies showed that the recombinant antigenic proteins of T. gondii may be an alternative source of antigens which are very useful for the serodiagnosis of toxoplasmosis. The article presents a review of current studies on the application and usefulness of different T. gondii recombinant antigens in serological tests for the diagnosis of human toxoplasmosis.
    Clinical and vaccine Immunology: CVI 06/2013; · 2.60 Impact Factor

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