A Bivalent Typhoid Live Vector Vaccine Expressing both Chromosome- and Plasmid-Encoded Yersinia pestis Antigens Fully Protects against Murine Lethal Pulmonary Plague Infection

Abstract

Live attenuated bacteria hold great promise as multivalent mucosal vaccines against a variety of pathogens. A major challenge of this approach has been the successful delivery of sufficient amounts of vaccine antigens to adequately prime the immune system without overattenuating the live vaccine. Here we used a live attenuated Salmonella enterica serovar Typhi strain to create a bivalent mucosal plague vaccine that produces both the protective F1 capsular antigen of Yersinia pestis and the LcrV protein required for secretion of virulence effector proteins. To reduce the metabolic burden associated with the coexpression of F1 and LcrV within the live vector, we balanced expression of both antigens by combining plasmid-based expression of F1 with chromosomal expression of LcrV from three independent loci. The immunogenicity and protective efficacy of this novel vaccine were assessed in mice by using a heterologous prime-boost immunization strategy and compared to those of a conventional strain in which F1 and LcrV were expressed from a single low-copy-number plasmid. The serum antibody responses to lipopolysaccharide (LPS) induced by the optimized bivalent vaccine were indistinguishable from those elicited by the parent strain, suggesting an adequate immunogenic capacity maintained through preservation of bacterial fitness; in contrast, LPS titers were 10-fold lower in mice immunized with the conventional vaccine strain. Importantly, mice receiving the optimized bivalent vaccine were fully protected against lethal pulmonary challenge. These results demonstrate the feasibility of distributing foreign antigen expression across both chromosomal and plasmid locations within a single vaccine organism for induction of protective immunity.T he process of engineering live attenuated organisms for mucosal delivery of protective foreign antigens has become a sophisticated enterprise, with powerful improvements in expression technologies occurring over the past 3 decades (1-5). To date, the most straightforward implementation of such expression technologies has involved the use of multicopy plasmids. Plasmids have been engineered to encode nonantibiotic selection markers which confer stable maintenance of these plasmids, both in vitro and after vaccination, thereby promoting optimum expression of sufficient levels of antigen to elicit protective immunity (6-8). Antigen export systems have also been devised to export antigens out of the cytoplasm and either onto the cell surface or out into the surrounding milieu (9-11). Export of foreign antigens is now appreciated to improve immune responses, possibly by avoiding proteolytic degradation of antigens within the cytoplasm or periplasmic space of the vaccine organism (10,(12)(13)(14)(15)(16)(17).However, there can be additional pitfalls introduced by stabilized expression plasmids. Sustained production of large amounts of foreign antigen can impose a metabolic burden upon the vaccine that overattenuates the strain and results in reduced immunogenicity (1,(18)(19)(20)(21)(22). T...

B cells and antibodies in the defense against Mycobacterium tuberculosis infection

“…For example, intraperitoneal injection of monoclonal antibody targeting a lipopolysaccharide of the Q fever producing facultative intracellular organism Coxiella burnetii has shown to protect mice against aerosolized infection . Also in accordance with these notions, vaccines based on the induction of antibodies that promote phagocytosis of respiratory, intracellular pathogens such as Yersinia pestis and Francisella tularemia have recently been developed and shown protection against plague and tularemia . Furthermore, alveolar macrophages and epithelial cells produce several proteins of the classical and alternative pathways of complement , and complement proteins have been detected in the bronchoalveolar lavage fluid from various different mammalian species including humans .…”

Plague Vaccines: Status and Future

“…The serum antibody responses to LPS induced by the optimized bivalent vaccine were indistinguishable from those elicited by the parent strain, suggesting adequate immunogenic capacity maintained through preservation of bacterial fitness. Importantly, mice receiving the optimized bivalent vaccine were fully protected against lethal pulmonary challenge [178]. …”

“…Salmonella -based vaccine administration is needle-free and easier and less expensive to manufacture (~10 cents/dose for human vaccines) than subunit vaccines. Several groups are endeavored to develop a vastly improved array of means to enhance the safety, efficacy and utility of Salmonella antigen delivery technologies [178, 271, 272]. Use of the Salmonella vector system for delivery of multiple Yersinia antigens also has good prospects against plague.…”

Animal Models for Salmonellosis: Applications in Vaccine Research

“…Finally, these early experiments in mice may allow researchers to characterize correlates of protection, allowing them to establish whether the vaccine could be effective in humans. Many Salmonella vaccine candidates have been tested in this model, including whole killed, live attenuated, subunit, conjugate, and Salmonella live vector vaccines (28,36,(53)(54)(55).…”