Conversion of RpoS ⁻ Attenuated Salmonella enterica Serovar Typhi Vaccine Strains to RpoS ⁺ Improves Their Resistance to Host Defense Barriers

Abstract

The vast majority of live attenuated typhoid vaccines are constructed from the <i>Salmonella enterica</i> serovar Typhi strain Ty2, which is devoid of a functioning alternative sigma factor, RpoS, due to the presence of a frameshift mutation. RpoS is a specialized sigma factor that plays an important role in the general stress response of a number of Gram-negative organisms, including <i>Salmonella</i>. Previous studies have demonstrated that this sigma factor is necessary for survival following exposure to acid, hydrogen peroxide, nutrient-limiting conditions, and starvation. In addition, studies with <i>Salmonella enterica</i> serovar Typhimurium and the mouse model of typhoid fever have shown that RpoS is important in colonization and survival within the infected murine host. We converted 4 clinically studied candidate typhoid vaccine strains derived from Ty2 [CVD908<i>-htrA</i>, Ty800, and χ9639(pYA3493)] and the licensed live typhoid vaccine Ty21a (also derived from Ty2) to RpoS⁺ and compared their abilities to withstand environmental stresses that may be encountered within the host to those of the RpoS^- parent strains. The results of our study indicate that strains that contain a functional RpoS were better able to survive following stress and that they would be ideal for further development as safe, effective vaccines to prevent <i>S</i>. Typhi infections or as vectors in recombinant attenuated <i>Salmonella</i> vaccines (RASVs) designed to protect against other infectious disease agents in humans. The <i>S</i>. Typhi strains constructed and described here will be made freely available upon request, as will the suicide vector used to convert <i>rpoS</i> mutants to RpoS⁺. <b>IMPORTANCE</b> Recombinant attenuated <i>Salmonella</i> vaccines (RASVs) represent a unique prevention strategy to combating infectious disease because they utilize the ability of <i>Salmonella</i> to invade and colonize deep effector lymphoid tissues and deliver hetero- and homologous derived antigens at the lowest immunizing dose. Our recent clinical trial in human volunteers indicated that an RpoS⁺ derivative of Ty2 was better at inducing immune responses than its RpoS^- counterpart. In this study, we demonstrate that a functional RpoS allele is beneficial for developing effective live attenuated vaccines against <i>S</i>. Typhi or in using <i>S</i>. Typhi as a recombinant attenuated vaccine vector to deliver other protective antigens.