To identify antigens of spp. that are potentially involved in stimulating a protective T-cell-mediated immune response, previous studies identified 10 clones from a 2308 genomic library with primed lymphocytes as probes. One selected positive clone (182) contained an insert of 1.2 kb which was identified, sequenced and characterised. The deduced amino acid sequence of the open reading frame (ORF) revealed 82% and 81% identity to the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) enzymes from and , respectively. Southern blot analysis demonstrated that the gene is present in only one copy in the genome. GAPDH was then expressed in as a fusion protein with the maltose-binding protein (MBP). To demonstrate the functional activity of GAPDH, mutants were transformed with a pMAL- construct. Genetic complementation was achieved and as a result mutants were able to grow on glucose or other carbon source medium. The humoral and cellular immune responses to the recombinant (r) GAPDH were characterised. In Western blots, sera from naturally infected cattle and sheep showed antibody reactivity against rGAPDH. In response to in-vitro stimulation by rGAPDH, splenocytes from mice vaccinated with rGAPDH or S19 were able to produce γ-interferon and tumour necrosis factor-α but not interleukin (IL)-4. Furthermore, associated with murine gene in a DNA vaccine formulation partially protected mice against experimental infection.


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