DEVELOPMENT OF Salmonella typhi Ty21a AS A POTENTIAL ORAL VACCINE AGAINST TUBERCULOSIS: SURFACE DISPLAY AND DNA VACCINE CARRIER OF A SYNTHETIC MULTI-EPITOPE MYCOBACTERIAL GENE by MOHAMMED ABDEL AZIZ AHMAD SARHAN

Despite the discovery of the causative agent of tuberculosis (TB), Mycobacterium tuberculosis, more than 120 years ago TB remains a major worldwide health problem. Currently, the attenuated strain of M. bovis, Bacille Calmette-Güerin (BCG) is the only vaccine available against TB. Although BCG is the world's most widely used vaccine, its protective value as an anti-TB vaccine for adults in certain areas of the world, has been shown to be low or even non-existent. Thus there is general agreement that new novel vaccines are required for TB control and prevention especially in developing countries. In this study, the use of the live attenuated typhoid vaccine, S. typhi Ty21a, for development as candidate vaccines against TB was explored in which the organism was utilized in a surface display system as well as a carrier of a DNA vaccine. In the surface display approach, a surface display expression system was developed by the construction of a synthetic gene coding for the N-terminal of the ice nucleation protein (Inak-n) from Pseudomonas syringae using a method called assembly polymerase chain reaction (PCR). In this method, the Inak-n gene was assembled from 34 overlapping chemically synthesized oligonucleotides in a single step and amplified by PCR using specific cloning primers. The gene was cloned into the pCR2.1-TOPO vector to create a recombinant plasmid designated as pMSInak. Cloning of a previously constructed 0.82kb synthetic gene known as VacII [which contained selected T cell epitopes of several M. tuberculosis genes namely ESAT6, MTP40, 38 kDa and MPT64 and further modified to include six consecutive histidine