Screening and identification of novel B cell epitopes of Toxoplasma gondii SAG1
© Wang et al.; licensee BioMed Central Ltd. 2013
Received: 4 January 2013
Accepted: 26 April 2013
Published: 30 April 2013
The identification of protein epitopes is useful for diagnostic purposes and for the development of peptide vaccines. In this study, the epitopes of Toxoplasma gondii SAG1 were identified using synthetic peptide techniques with the aid of bioinformatics.
Eleven peptides derived from T. gondii SAG1 were assessed by ELISA using pig sera from different time points after infection. Four (PS4, PS6, PS10 and PS11), out of the eleven peptides tested were recognized by all sera. Then, shorter peptides that were derived from PS4, PS6, PS10 and PS11 were predicted using bioinformatics and tested by experimentation. Four out of nine shorter peptides were identified successfully (amino acids 106–120, 166–180, 289–300 and 313–332).
We have precisely located the epitopes of T. gondii SAG1 using pig sera collected at different time points after infection. The identified epitopes may be useful for the further study of epitope-based vaccines and diagnostic reagents.
Toxoplasma gondii is an obligate intracellular parasite that infects a variety of mammals and birds, causing toxoplasmosis [1, 2]. T. gondii is an important food-borne parasite, and the primary route of transmission from animals to humans is through the consumption of infected meat [3, 4]. In some countries, pork is the most common meat consumed, and some ethnic groups consume raw pork; thus, pigs are considered to be the primary source of human infection with T. gondii. In addition, toxoplasmosis is a source of significant economic losses for swine farmers . Therefore, the development of effective diagnostic reagents or vaccines for controlling this infection is required. Attempts to develop a peptide-based vaccine for T. gondii have focused on SAG1 and shown encouraging results . Furthermore, the multiepitope antigen is one of the most promising antigens for the serodiagnosis of toxoplasmosis. Thus, it is very important to determine the precise sequences against which effective immune responses are directed. SAG1 epitopes have been studied by different research groups [8–10]. However, it is still unclear which SAG1 peptides are recognized by antibodies from pigs infected with T. gondii. Therefore, B cell epitopes of SAG1 were analyzed using a synthetic peptide technique in combination with software-based prediction.
A total of 51 T. gondii-positive sera, which had been previously collected from experimentally infected pigs by our lab, were investigated. Twelve pig serum samples were collected at the time of presentation of clinical symptoms (G1); 18 follow-up specimens were taken on days 14 to 35 after the onset of symptoms (G2), and 21 further serum samples were taken on days 60 to 120 after the onset of symptoms (G3). Toxoplasma IgM and IgG antibodies were confirmed by T. gondii lysate antigen-ELISA. The serum samples in G1 and G2 were positive for IgM and IgG against T. gondii. The serum samples in G3 were only positive for IgG against T. gondii. Ten sera that were negative for T. gondii IgM and IgG were used as controls. The experimental protocol was approved by the Ethical Committee of the Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences.
Sequences of synthesized peptides
Start and end position
Precise definition of the epitopes
Conformational epitope selection needs to determine the tertiary structure of antigen in order to identify antibody interacting residues in antigen. Experimental techniques such as crystallography are expensive and time consuming. Thus, our work focuses on linear epitope selection. In this study, we have located the epitopes of T. gondii SAG1 to a shorter sequence than had been identified previously. The identified epitopes will be useful in vaccine and diagnostic reagent design.
This investigation was supported by grants from the National Special Research Programs for Non-profit Trades (Agriculture) (200903036–02) and NBCITS, MOA (CARS-38).
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