Characterization of O-antigen delivered by Generalized Modules for Membrane Antigens (GMMA) vaccine candidates against nontyphoidal Salmonella.

Abstract Invasive nontyphoidal Salmonella disease (iNTS) is a leading cause of death and morbidity in Africa. The most common pathogens are Salmonella entericaserovars Typhimurium and Enteritidis. The O- antigenportion of their lipopolysaccharideis a target of protective immunity and vaccines targeting O- antigenare currently in development. Here we investigate the use of Generalized Modules for Membrane Antigens(GMMA) as delivery system for S . Typhimurium and S . Enteritidis O- antigen. Gram-negative bacteria naturally shed outer membrane in a blebbing process. By deletion of the tolR gene, the level of shedding was greatly enhanced. Further genetic modifications were introduced into the GMMA-producing strains in order to reduce reactogenicity, by detoxifying the lipid A moiety of lipopolysaccharide. We found that genetic mutations can impact on expression of O- antigenchains. All S. Enteritidis GMMA characterized had an O- antigento protein w/w ratio higher than 0.6, while the ratio was 0.7 for S. Typhimurium Δ tolR GMMA, but decreased to less than 0.1 when further mutations for lipid A detoxification were introduced. Changes were also observed in O- antigenchain length and level and/or position of O-acetylation. When tested in mice, the GMMA induced high levels of anti-O- antigen-specific IgG functional antibodies, despite variation in density and O- antigenstructural modifications. In conclusion, simplicity of manufacturing process and low costs of production, coupled with encouraging immunogenicity data, make GMMA an attractive strategy to further investigate for the development of a vaccine against iNTS.