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.

幽门螺杆菌（Helicobacter pylori）是一种全球流行的感染性病原，与慢性胃炎密切相关，甚至可诱发胃癌的发生。随着抗生素耐药性问题日益凸显，科研人员开始探索更优化的疫苗设计策略，以根除幽门螺杆菌的定植感染。然而当前主流的疫苗策略多采用单一幽门螺杆菌抗原，其应用潜力尚未得到充分发掘。外膜囊泡（Outer Membrane Vesicles, OMVs）是一类极具潜力的疫苗递送平台，因其可同时递送多种抗原。本研究借助血红蛋白蛋白酶（hemoglobin protease, Hbp）自转运系统，将三种关键的幽门螺杆菌抗原蛋白（UreB、CagA与VacA）展示于肠炎沙门氏菌鼠伤寒血清型（Salmonella enterica serovar Typhimurium）突变菌株来源的外膜囊泡表面。通过多组敲除策略筛选发现，从ΔrfbP ΔfliC ΔfljB ΔompA突变菌株中分离得到的外膜囊泡，可显著提升免疫球蛋白G（IgG）与免疫球蛋白A（IgA）的机体水平，并有效激活辅助性T细胞1型（Th1）及17型（Th17）偏向的细胞免疫应答，该应答在抵御幽门螺杆菌感染中发挥关键保护作用。后续本研究以该ΔrfbP ΔfliC ΔfljB ΔompA突变菌株来源的外膜囊泡为递送载体，搭载不同组合的幽门螺杆菌抗原。小鼠模型中的抗体与细胞因子水平检测及攻毒实验结果表明，共递送UreB与CagA可有效抵御幽门螺杆菌感染，并触发抗原特异性T细胞应答。综上，以ΔrfbP ΔfliC ΔfljB ΔompA突变菌株来源的外膜囊泡为递送载体，借助Hbp自转运系统导入UreB与CagA作为抗原，将为幽门螺杆菌感染的防控提供重要助力。