Effect of expression level on immune responses to recombinant oral Salmonella enterica serovar Typhimurium vaccines

Live, attenuated Salmonella has been used to express heterologous antigens for development of oral vaccines. Often, expression must be regulated because of deleterious effects on the Salmonella vector. The effect of varying expression levels on immune response parameters has not been well defined. In this study we introduced mutations in the -10 region of the trc promoter in the expression plasmid to generate series of vaccine strains with different levels of expression of a model antigen, the hemagglutinin HagB from Porphyromonas gingivalis. There was no difference in growth rates of the Salmonella vaccine strains containing the wild-type, the mutant plasmids or the empty expression vector. The primary IgG response in serum in mice orally immunized with the wild-type strain peaked 3-4 weeks earlier than the intermediate expression level strains, suggesting that high expression levels may favor an earlier response. While there was a trend for anti-HagB recall responses to correlate with higher expression level, the peak levels were not significantly different even for expression levels as low as 33% of wild-type. A similar trend in terms of response level was seen with serum and salivary IgA. The subclass of the IgG response was predominately IgG2a regardless of expression level, consistent with a Th1 response. These data suggest that isotype distribution, immune response level and T helper cell profile are largely unaffected over a wide range of expression levels.