Outer membrane vesicles from genetically engineered Salmonella enterica serovar Typhimurium presenting Helicobacter pylori antigens UreB and CagA induce protection against Helicobacter pylori infection in mice

<em>Helicobacter pylori</em> is a globally prevalent infection that is highly related to chronic gastritis and even causes the development of gastric carcinoma. With the rapid increase of antibiotic resistance, scientists have begun to search for better vaccine design strategies to eradicate <em>H. pylori</em> colonization. However, while current strategies prefer to formulate vaccines with a single <em>H. pylori</em> antigen, their potential has not yet been fully realized. Outer membrane vesicles (OMVs) are a potential vaccine platform since they could deliver multiple antigens. In this study, we engineered three crucial <em>H. pylori</em> antigen proteins (UreB, CagA, and VacA) onto the surface of OMVs derived from <em>Salmonella</em> <em>enterica serovar </em>Typhimurium mutant strains using the hemoglobin protease (Hbp) autotransporter system. In various knockout strategies, we found that OMVs isolated from the Δ<em>rfbP</em> Δ<em>fliC</em> Δ<em>fljB</em> Δ<em>ompA</em> mutants could cause distinct increases in immunoglobulin G (IgG) and A (IgA) levels and effectively trigger T helper 1- and 17-biased cellular immune responses, which perform a vital role in protecting against<em> H. pylori</em>. Next, OMVs derived from Δ<em>rfbP</em> Δ<em>fliC</em> Δ<em>fljB</em> Δ<em>ompA</em> mutants were used as a vector to deliver different combinations of <em>H. pylori</em> antigens. The antibody and cytokine levels and challenge experiments in mice model indicated that co-delivering UreB and CagA could protect against <em>H. pylori</em> and antigen-specific T cell responses. In summary, adopting OMVs derived from the Δ<em>rfbP</em> Δ<em>fliC</em> Δ<em>fljB</em> Δ<em>ompA</em> mutant strain as the vector while importing UreB and CagA as antigens using the Hbp autotransporter system would greatly benefit controlling<em> H. pylori</em> infection.