Outer membrane protein size and LPS O-antigen define protective antibody targeting to the Salmonella surface (1)
Antibodies to the Gram-negative bacterial surface, induced after natural infection or vaccination, save millions of lives each year, and typically function before disease develops, reducing the need to use antibiotics and the development of antimicrobial resistance. The only licensed single-antigen vaccines against diseases caused by such bacteria are based on capsular polysaccharides. In Gram-negative bacteria, polysaccharide O-antigen (O-Ag) made up of varying numbers of repeats, sometimes > 100 in number, restricts access of molecules to the bacterial surface. The cell-surface protein, IgG antibodies to the porin protein OmpD can offer cell-dependent or complement-dependent protection, and not all IgG isotypes offer equivalent protection correlating to the level of access of antibodies to bind OmpD in the presence of O-Ag.
Wild-type (WT) mice were immunized twice with 20 μg of purified STmOmpD. The induced anti-STmOmpD IgG bound to the surface of two different isolates of STm: the model laboratory strain SL1344 and a representative invasive African isolate of STm, D23580, which was originally isolated from an African patient. anti-STmOmpD antibodies enhanced complement-dependent killing, whereas sera from control mice did not.
STm and SEn in which OmpD differs by a single amino acid, a substitution of an alanine in STmOmpD at position 263 to a serine in SEn. Immunization with STmOmpD induced statistically significant protection in both groups. However, while there was a near-100-fold reduction in immunized mice challenged with STm, the reduction in bacterial numbers in SEn-challenged mice was less than fivefold.
Binding of anti-STmOmpD IgG to O-Ag-expressing STm or O-Ag-deficient STm was similar, consistent with the antibody having a solely proteinaceous target. In contrast, when the coating antigen used was whole WT SEn, then antibody binding was dramatically lower than that observed for the different STm strains. Anti-STmOmpD antibody was bactericidal for STm, but not SEn. Most IgG that binds OmpD on O-Ag-expressing bacteria targets a limited number of variable proteinaceous epitopes that include A263S, whereas O-Ag occludes binding to the larger number of epitopes conserved between the two proteins.
Nevertheless, antibodies to purified OmpA bound only weakly to intact bacteria, and did not induce protective responses. Therefore, despite being a target of naturally induced antibodies, this protein in STm and SEn is not under immune pressure, and does not induce protective responses. Sequence variation in the surface-exposed loops of other outer-membrane proteins, such as the monomeric proteins OmpX and OmpW, and the trimeric porin OmpC is similar to the data. OmpX and OmpW are identical between STm and SEn, but there are differences in OmpC, which are conserved between STm and SEn isolates, respectively, and thus may reflect some association with O-Ag usage. There is no single surface protein antigen subunit vaccine for Gram-negative bacteria and if a capsule antigen is not used in the vaccine, then they increase considerably in their complexity. One reason for this increased complexity may be the influence of LPS/LOS O-Ag on the accessibility of antibody to the component antigens, meaning that greater antigen coverage is needed to ensure protection. In contrast, since capsules are typically located on the outermost surface of the bacterium, it may mean there is less impediment to antibody binding and less need to include multiple antigens in a vaccine.