Construction and characterisation of Salmonella typhimurium aroA simultaneously expressing the five pertussis toxin subunits.

Pertussis toxin (PT) is a major protective antigen of Bordetella pertussis, the causative agent of whooping cough. Current vaccines against whooping cough are administered parenterally, and generate a systemic immune response. An alternative to this approach is to stimulate mucosal and systemic immune responses by oral immunisation with live vaccine strains of Salmonella spp. We have individually expressed the five PT subunits in Salmonella typhimurium aroA and subsequently expressed a synthetic pertussis toxin operon (pDP16), via the use of a temperature inducible expression system. In S. typhimurium aroA containing individual subunit expression plasmids the S5 subunit was found to be completely processed, while all other subunits were partially processed. In S. typhimurium aroA (pDP16) S5 was completely processed, S1 was partially processed, whilst S2, S3 and S4 remained unprocessed. Induction of the synthetic PT operon decreased the in vitro invasiveness of S. typhimurium aroA, compared to vector-only and plasmid-less strains. Following oral immunisation of Balb/c mice with S. typhimurium aroA (pDP16), statistically significant IgG (p < 0.05) specific to PT was detected in the serum of mice. Nonetheless, despite a statistically significant anti-PT serum antibody response in immunised mice, vaccination with S. typhimurium aroA (pDP16) failed to protect mice from virulent challenge.