# The JMU-SalVac-System: A Novel, Versatile Approach to Oral Live Vaccine Development

## TL;DR

Researchers developed a new oral vaccine system using a modified Salmonella strain to safely and effectively deliver antigens for mucosal immunization.

## Contribution

A novel balanced-lethal system (BLS) ensures plasmid stability in an oral Ty21a-based vaccine platform for human use.

## Key findings

- The JMU-SalVac-system uses a ΔtyrS/tyrS+-based BLS to stabilize antigen delivery plasmids in Ty21a.
- Cytosolic and secreted model antigens (mRFP and CTB) were successfully expressed in vaccine strains.
- Promoters in pSalVac-plasmids were induced in primary macrophages, demonstrating in vivo functionality.

## Abstract

Salmonella enterica Serovar Typhi Ty21a (Ty21a) is the only licensed oral vaccine against typhoid fever. Due to its excellent safety profile, it has been used as a promising vector strain for the expression of heterologous antigens for mucosal immunization. As the efficacy of any bacterial live vector vaccine correlates with its ability to express and present sufficient antigen, the genes for antigen expression are traditionally located on plasmids with antibiotic resistance genes for stabilization. However, for use in humans, antibiotic selection of plasmids is not applicable, leading to segregational loss of the antigen-producing plasmid. Therefore, we developed an oral Ty21a-based vaccine platform technology, the JMU-SalVac-system (Julius-Maximilians-Universität Würzburg) in which the antigen delivery plasmids (pSalVac-plasmid-series) are stabilized by a ΔtyrS/tyrS+-based balanced-lethal system (BLS). The system is made up of the chromosomal knockout of the essential tyrosyl-tRNA-synthetase gene (tyrS) and the in trans complementation of tyrS on the pSalVac-plasmid. Further novel functional features of the pSalVac-plasmids are the presence of two different expression cassettes for the expression of protein antigens. In this study, we present the construction of vaccine strains with BLS plasmids for antigen expression. The expression of cytosolic and secreted mRFP and cholera toxin subunit B (CTB) proteins as model antigens is used to demonstrate the versatility of the approach. As proof of concept, we show the induction of previously described in vivo inducible promoters cloned into pSalVac-plasmids during infection of primary macrophages and demonstrate the expression of model vaccine antigens in these relevant human target cells. Therefore, antigen delivery strains developed with the JMU-SalVac technology are promising, safe and stable vaccine strains to be used against mucosal infections in humans.