Document Type : Original Article
Department of Microbiology, Karaj Branch, Islamic Azad University, Karaj, Iran
Faculty of Chemistry and Chemical Engineering, Malek Ashtar University of Technology, Iran
Department of Biology, Faculty of Science, Imam-Hossein University, Tehran, Iran
Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran
Introduction: Shigella dysenteriae with a low infectious dose and its strains that are resistant to antibiotics is considered a health problem all over the world. IpaD as an important protein in the shigella type 3 secretion systems can be a convenient target for designing recombinant subunit vaccines against the bacteria due to its immunogenic properties. The present study aimed to evaluate the immunogenicity of a recombinant protein containing immunogenic regions of IpaD as a subunit recombinant vaccine candidate against Shigella dysenteriae.
Materials and Methods: The gene encoding immunogenic segments of ipaD was sub-cloned into pET28a expression vector. The new plasmid (pET-IpaD) was transformed into E. coli strain Rosetta (DE3). The recombinant protein was then expressed and purified using affinity chromatography and confirmed by western blotting. The guinea pigs were then immunized with purified protein and the antibody titer and specificity of the sera were analyzed by ELISA. Finally, an animal challenge was performed using the Sereny test.
Results: According to the designed pET-IpaD plasmid, the expression of recombinant protein in E. coli caused the production of a recombinant protein with 22 kDa molecular weight, and the western blot technique indicated the reaction of recombinant protein with anti-histidine monoclonal antibody. The yield of the purified protein from the culture medium was estimated at about 0.57 mg/ml and the immunogenic effect of the produced protein was determined by using in vitro and in vivo studies.
Conclusions: According to the findings of the present study, it can be concluded that the recombinant produced IpaD is a perfect peptide vaccine candidate for the development of a recombinant vaccine against Shigella dysenteriae.