Expression and Immunogenicity of Hemagglutinin A from Porphyromonas gingivalis in an Avirulent Salmonella enterica Serovar Typhimurium Vaccine Strain

Abstract

Porphyromonas gingivalis is a major etiologic agent of periodontitis, a chronic inflammatory disease that ultimately results in the loss of the supporting tissues of the teeth. Previous work has demonstrated the usefulness of avirulent Salmonella enterica serovar Typhimurium strains as antigen delivery systems for protective antigens of pathogens that colonize or cross mucosal surfaces. In this study, we constructed and characterized a recombinant S. enterica serovar Typhimurium avirulent vaccine strain which expresses hemagglutinin A and carries no antibiotic resistance markers. HagA, a major virulence-associated surface protein, is a potentially useful immunogen that contains an antigenic epitope which, in humans, elicits an immune response that is protective against subsequent colonization by P. gingivalis. The hagA gene, including its promoter, was cloned into a balanced-lethal Salmonella vector and transferred to the vaccine strain. Heterologous expression of HagA was demonstrated in both Escherichia coli JM109 and S. enterica serovar Typhimurium vaccine strain 4072. The HagA epitope was present in its native configuration as determined by immunochemistry and immunoelectron microscopy. Purified recombinant HagA was recognized by sera from mice immunized with the S. enterica serovar Typhimurium vaccine strain. The HagA-specific antigen of the vaccine was also found to be recognized by serum from a periodontal patient. This vaccine strain, which expresses the functional hemagglutinin protein, induces a humoral immune response against HagA and may be useful for developing a protective vaccine against periodontal diseases associated with P. gingivalis.

Nasal Immunization with a Fusion Protein Consisting of the Hemagglutinin A Antigenic Region and the Maltose-Binding Protein Elicits CD11c+CD8+Dendritic Cells for Induced Long-Term Protective Immunity

“…A previous study confirmed by Western immunoblotting that oral immunization with recombinant avirulent Salmonella enterica serovar Typhimurium expressing HagA induces HagAspecific serum antibody responses (30). In addition, a hen egg yolk antibody (IgY) against a truncated HagA protein has been developed (53).…”

“…Multiple hemagglutinin genes have been cloned from P. gingivalis by functional screening (35,42,44,49). Among these, hemagglutinin A (HagA) has been thought to contain the functional domain of hemagglutinin and to be a potentially useful immunogen that elicits a protective immune response against subsequent colonization by P. gingivalis (30). The hagA gene is 7,887 bp long and encodes a protein of 2,628 amino acids, with a molecular mass of 283.3 kDa (17).…”

Both the unique and repeat regions of the Porphyromonas gingivalis hemagglutin A are involved in adhesion and invasion of host cells

“…E. coli pEKS5 carries the same insert in expression vector therefore we expect to have similar localization. Based on these data, it was not surprising to observe, using EM, that the (entire) HagA, expressed in S. enterica , was also surface-associated [40]. …”

“…A vaccine strain of Salmonella enterica serovar Typhimurium χ4072 ( S. enterica pNM1.1), which expresses the functional hemagglutinin A of P. gingivalis , and it respective control strain harboring an empty vector were described in a previous study and grown in LB broth or plates [40]. A diagram of S. enterica pNM1.1 insert can be found in Fig.…”

“…Due to its importance, HagA was expressed in an immunogenic form in a Salmonella enterica serovar Typhimurium avirulent vaccine strain. This recombinant major virulence associated surface protein was recognized by serum from a periodontal patient [40]. This vaccine strain could be used to develop a protective vaccine against P. gingivalis infection.…”