A novel DNA vaccine against toxoplasmosis induces sporozoite specific protective immune response through nonapoptotic cells
Abstract number: O151
Gulce Iz S., Doskaya M., Caner A., Degirmenci A., Rodriguez F., Deliloglu Gurhan I., Guruz Y.
Objectives:Toxoplasma gondii is an obligate intracellular parasite infecting all warm-blooded animals, including humans and causes serious clinical presentations. There is no 100% effective drug to treat toxoplasmosis. Development of a vaccine, which can prevent the consequences of acute infection, is an attractive alternative.
Since 1990s, vaccination strategies against toxoplasmosis mainly used DNA vaccine, purified recombinant proteins showed variable protection because vaccine candidate antigens were almost always selected randomly and targeted tachyzoite form which invades the host cell abruptly and immediately embeds itself in a protective parasitophorous vacuole which keeps away the host immune response.
Currently, there is substantial evidence about T. gondii oocysts (contains sporozoites) as being the cause of water related outbreaks. After classification of T. gondii oocysts in category B bioterrorism agents as a water safety threat, the demand for a protective vaccine against toxoplasmosis has increased.
The present study aims to generate first time a DNA vaccine containing a sporozoite specific surface antigen "SporoSAG" to block the sporozoites as they are released from the oocysts in the intestine. To increase the efficacy of the vaccine, antigen specific-CD8 response inducing anti-apoptotic Bcl-xL gene was inserted to the dual expression DNA vaccine.
Methods: During the construction of the dual expression DNA vaccine, SporoSAG was inserted after CMV promoter and anti-apoptotic protein Bcl-xL were inserted in frame with EGFP after IRES (pIRES2EGFP-SporoSAG-Bcl-xL). In vitro transfection of BHK-21 cells was performed to show the expression of SporoSAG and Bcl-xL. The functionality of Bcl-xL was demonstrated by Casp3 flow cytometry. Humoral and cellular immune response (CD4/CD8, IFN-g) were analyzed from sera and spleen cells of vaccinated BALB/c mice by western blot and flow cytometry using purified recombinant SporoSAG protein (Figure 1).
Results and Conclusion: Western blot and fluorescence microscopy analyses of pIRES2EGFP-SporoSAG-Bcl-xL transfected BHK-21 cells showed SporoSAG and Bcl-xL expression. Casp3 analyses rationalized Bcl-xL expression that impedes apoptotic cell death. Analysis of sera obtained from vaccinated mice showed anti-SporoSAG antibody response compared to controls. Cellular immune response analyses showed increased CD8 and IFN-g response compared to controls indicative of protection against toxoplasmosis.
|Session name:||Abstracts 20th European Congress of Clinical Microbiology and Infectious Diseases|
|Location:||Vienna, Austria, 10 - 13 April 2010|
|Back to top|